Friday, September 24, 2021

NORWAY CONFIRMS ATYPICAL CWD TSE Prion DEER IN ETNE

NORWAY ATYPICAL CWD TSE PRION

NORWAY CONFIRMS ATYPICAL SCRAP DISEASE ON DEER IN ETNE

24.09.2021 BY: MATTILSYNET

Final test results show that the deer that was killed while hunting in Etne had atypical scrapie. This is a variant of scrapie that we consider non-contagious. - This is good news, says Anne Marie Jahr, section manager for animal health in the Norwegian Food Safety Authority.

Confirms atypical scrapie on deer in Etne There was a suspicion of scrapie in a deer that was killed in Etne on 11 September. Final test results show that this is the variant that is considered non-contagious. Stock Photo: Svein-Håkon Lorentsen, NINA - It is not dramatic, and we expect some such cases a year. Had this been the classic variant, which is contagious, it would have been serious. This is an important reminder of how central hunters are in the mapping of this disease. Samples from animals that are killed during hunting are important, and I would like to thank those who contribute to the survey, says Jahr.

In order to get even more samples in the area, we have now incorporated Sauda municipality into the Norwegian Food Safety Authority's national mapping program for scrapie. The other neighboring municipalities to Etne are already included in the program. This is done to get a better overview of the area.

Two variants of scrapie There are two variants of scrapie, an infectious variant (classic) and another variant that can probably occur spontaneously in older animals (atypical). Classic scrapie has previously been found in wild reindeer in Nordfjella and on the Hardangervidda. Atypical scrapie has been found on elk and deer in various parts of the country.

Report if you see sick or dead deer

Since the start in 2016, more than 130,000 deer have been tested for scrapie. The Norwegian Food Safety Authority encourages everyone who travels in forests and fields, and who sees sick or dead deer, to report to the Norwegian Food Safety Authority. Symptoms of scrapie are weight loss, frequent urination and abnormal behavior, e.g. animals that do not shy away from humans.

Read more about the mapping program for scrapie

Read the Norwegian Food Safety Authority's press release


***> This is a variant of scrapie that we consider non-contagious. 

***> Atypical/Nor98 scrapie occurs in sheep that tend to be resistant to classical scrapie and it is thought to occur spontaneously.

 ***> Unlike classical scrapie (CS), AS is thought to be a spontaneously occurring disease [1–3].

***> It typically affects a single older sheep within a flock, and cases of AS are sporadic and isolated suggesting that natural transmission is unlikely.

***> Several experiments have demonstrated the ability of the AS agent to transmit within the natural host after intracranial inoculation [14–16]. 

***> One study found that the AS agent could transmit after a high oral dose of AS brain homogenate [17]. 

***> Nonetheless, AS is still considered unlikely to transmit under field conditions; therefore, eradication and surveillance programs for CS have allowed exceptions for AS. 

I disagree, and this is a foolish move taking this stance, because Science has shown us that the atypical scrapie IS TRANSMISSIBLE!

EFSA SAYS ATYPICAL SCRAPIE ROUGHLY HAS 50 50 CHANCE ATYPICAL SCRAPIE IS CONTAGIOUS, AS NON-CONTAGIOUS!

TO CONTINUE THIS CHARADE, THAT ''variant of scrapie that we consider non-contagious'' and ''AS is thought to be a spontaneously occurring disease'', WILL ONLY CONTINUE TO SPREAD CWD TSE PRION, INCLUDING ATYPICAL SCRAPIE...TERRY

***Moreover, sporadic disease has never been observed in breeding colonies or primate research laboratories, most notably among hundreds of animals over several decades of study at the National Institutes of Health25, and in nearly twenty older animals continuously housed in our own facility.***

Even if the prevailing view is that sporadic CJD is due to the spontaneous formation of CJD prions, it remains possible that its apparent sporadic nature may, at least in part, result from our limited capacity to identify an environmental origin.


THURSDAY, JULY 8, 2021

EFSA Scientific report on the analysis of the 2‐year compulsory intensified monitoring of atypical scrapie

***> AS is considered more likely (subjective probability range 50–66%) that AS is a non-contagious, rather than a contagious, disease.

ATYPICAL SCRAPIE ROUGHLY HAS 50 50 CHANCE ATYPICAL SCRAPIE IS CONTAGIOUS, AS NON-CONTAGIOUS, TAKE YOUR PICK, BUT I SAID IT LONG AGO WHEN USDA OIE ET AL MADE ATYPICAL SCRAPIE A LEGAL TRADING COMODITY, I SAID YOUR PUTTING THE CART BEFORE THE HORSE, AND THAT'S EXACTLY WHAT THEY DID, and it's called in Texas, TEXAS TSE PRION HOLDEM POKER, WHO'S ALL IN $$$

THURSDAY, JULY 8, 2021

EFSA Scientific report on the analysis of the 2‐year compulsory intensified monitoring of atypical scrapie


Experimental Oral Transmission of Atypical Scrapie to Sheep

Marion M. Simmons, S. Jo Moore,1 Timm Konold, Lisa Thurston, Linda A. Terry, Leigh Thorne, Richard Lockey, Chris Vickery, Stephen A.C. Hawkins, Melanie J. Chaplin, and John Spiropoulos 

To investigate the possibility of oral transmission of atypical scrapie in sheep and determine the distribution of infectivity in the animals’ peripheral tissues, we challenged neonatal lambs orally with atypical scrapie; they were then killed at 12 or 24 months. Screening test results were negative for disease-specifi c prion protein in all but 2 recipients; they had positive results for examination of brain, but negative for peripheral tissues. Infectivity of brain, distal ileum, and spleen from all animals was assessed in mouse bioassays; positive results were obtained from tissues that had negative results on screening. These fi ndings demonstrate that atypical scrapie can be transmitted orally and indicate that it has the potential for natural transmission and iatrogenic spread through animal feed. Detection of infectivity in tissues negative by current surveillance methods indicates that diagnostic sensitivity is suboptimal for atypical scrapie, and potentially infectious material may be able to pass into the human food chain.

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imals; the incubation periods of sheep orally infected with classical scrapie were signifi cantly shorter in sheep challenged at 14 days of age than those challenged at 6 months of age (31). If, however, oral transmission is only effective in such young animals, then fi eld exposure would most likely have to be through milk, which is known to be a highly effective route of transmission for classical scrapie (32). No data are currently available on the potential infectivity of milk from animals with atypical scrapie.

Successful oral transmission also raises questions regarding the pathogenesis of this form of disease. There must be passage of the infectious agent from the alimentary canal to the brain through one of several possible routes, most likely those that have been suggested and discussed in detail for other TSEs, for example, retrograde neuronal transportation either directly (33–35) or through lymphoid structures or hematogenously (36). Infectivity in the absence of readily demonstrable PrPSc has been reported (37–39), and although the mouse bioassay may detect evidence of disease in other tissues, these data may not be available for at least another 2 years. More protease-sensitive forms of PrPSc may be broken down more effi ciently within cells and thus do not accumulate in peripheral tissues (19), enabling atypical PrPSc to transit the digestive tract and disseminate through other systems in small amounts before accumulating detectably in the central nervous system.

Although we do not have epidemiologic evidence that supports the effi cient spread of disease in the fi eld, these data imply that disease is potentially transmissible under fi eld situations and that spread through animal feed may be possible if the current feed restrictions were to be relaxed. Additionally, almost no data are available on the potential for atypical scrapie to transmit to other food animal species, certainly by the oral route. However, work with transgenic mice has demonstrated the potential susceptibility of pigs, with the disturbing fi nding that the biochemical properties of the resulting PrPSc have changed on transmission (40). The implications of this observation for subsequent transmission and host target range are currently unknown. 

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A newly identified type of scrapie agent can naturally infect sheep with resistant PrP genotypes 

 Annick Le Dur*,?, Vincent Béringue*,?, Olivier Andréoletti?, Fabienne Reine*, Thanh Lan Laï*, Thierry Baron§, Bjørn Bratberg¶, Jean-Luc Vilotte?, Pierre Sarradin**, Sylvie L. Benestad¶, and Hubert Laude*,? +Author Affiliations 

*Virologie Immunologie Moléculaires and ?Génétique Biochimique et Cytogénétique, Institut National de la Recherche Agronomique, 78350 Jouy-en-Josas, France; ?Unité Mixte de Recherche, Institut National de la Recherche Agronomique-Ecole Nationale Vétérinaire de Toulouse, Interactions Hôte Agent Pathogène, 31066 Toulouse, France; §Agence Française de Sécurité Sanitaire des Aliments, Unité Agents Transmissibles Non Conventionnels, 69364 Lyon, France; **Pathologie Infectieuse et Immunologie, Institut National de la Recherche Agronomique, 37380 Nouzilly, France; and ¶Department of Pathology, National Veterinary Institute, 0033 Oslo, Norway 

***Edited by Stanley B. Prusiner, University of California, San Francisco, CA (received for review March 21, 2005) 

Abstract Scrapie in small ruminants belongs to transmissible spongiform encephalopathies (TSEs), or prion diseases, a family of fatal neurodegenerative disorders that affect humans and animals and can transmit within and between species by ingestion or inoculation. Conversion of the host-encoded prion protein (PrP), normal cellular PrP (PrPc), into a misfolded form, abnormal PrP (PrPSc), plays a key role in TSE transmission and pathogenesis. The intensified surveillance of scrapie in the European Union, together with the improvement of PrPSc detection techniques, has led to the discovery of a growing number of so-called atypical scrapie cases. These include clinical Nor98 cases first identified in Norwegian sheep on the basis of unusual pathological and PrPSc molecular features and "cases" that produced discordant responses in the rapid tests currently applied to the large-scale random screening of slaughtered or fallen animals. Worryingly, a substantial proportion of such cases involved sheep with PrP genotypes known until now to confer natural resistance to conventional scrapie. Here we report that both Nor98 and discordant cases, including three sheep homozygous for the resistant PrPARR allele (A136R154R171), efficiently transmitted the disease to transgenic mice expressing ovine PrP, and that they shared unique biological and biochemical features upon propagation in mice. *** These observations support the view that a truly infectious TSE agent, unrecognized until recently, infects sheep and goat flocks and may have important implications in terms of scrapie control and public health. 


ACCEPTED MANUSCRIPT

North American and Norwegian Chronic Wasting Disease prions exhibit different potential for interspecies transmission and zoonotic risk

Sandra Pritzkow, Damian Gorski, Frank Ramirez, Glenn C Telling, Sylvie L Benestad, Claudio Soto

The Journal of Infectious Diseases, jiab385, https://doi.org/10.1093/infdis/jiab385

Published: 24 July 2021 Article history Abstract

Chronic wasting disease (CWD) is a rapidly spreading prion disorder affecting various species of wild and captive cervids. The risk that CWD poses to co-habiting animals or more importantly to humans is largely unknown. 

In this study we investigated differences in the capacity of CWD isolates obtained from six different cervid species to induce prion conversion in vitro by PMCA. We define and quantify spillover and zoonotic potential indices as the efficiency by which CWD prions sustain prion generation in vitro at expenses of normal prion proteins from various mammals and human, respectively. 

Our data suggest that reindeer and red deer from Norway could be the most transmissible CWD prions to other mammals, whereas North American CWD prions were more prone to generate human prions in vitro. 

Our results suggest that Norway and North American CWD prions correspond to different strains with distinct spillover and zoonotic potentials.

prions, prion disease, chronic wasting disease, CWD, prion strains, PMCA, spillover potential, zoonotic potential, moose, reindeer, red deer, elk, white-tailed deer, mule deer

Topic: prion diseases prions reindeer wasting disease, chronic norwegian peritoneal mucinous carcinomatosis

Issue Section: Major Article


Chronic wasting disease: a cervid prion infection looming to spillover

Alicia Otero 1 2 3, Camilo Duque Velásquez 1 2, Judd Aiken 2 4, Debbie McKenzie 5 6

Affiliations expand

PMID: 34488900 DOI: 10.1186/s13567-021-00986-y



Transmission of the atypical/Nor98 scrapie agent to Suffolk sheep with VRQ/ARQ, ARQ/ARQ, and ARQ/ARR genotypes

Eric D. Cassmann,Najiba Mammadova,S. Jo Moore,Sylvie Benestad,Justin J. Greenlee 

Published: February 11, 2021https://doi.org/10.1371/journal.pone.0246503

Citation: Cassmann ED, Mammadova N, Moore SJ, Benestad S, Greenlee JJ (2021) Transmission of the atypical/Nor98 scrapie agent to Suffolk sheep with VRQ/ARQ, ARQ/ARQ, and ARQ/ARR genotypes. PLoS ONE 16(2): e0246503. https://doi.org/10.1371/journal.pone.0246503

Editor: Gianluigi Zanusso, University of Verona, ITALY

Received: November 24, 2020; Accepted: January 21, 2021; Published: February 11, 2021

Abstract

Scrapie is a transmissible spongiform encephalopathy that occurs in sheep. Atypical/Nor98 scrapie occurs in sheep that tend to be resistant to classical scrapie and it is thought to occur spontaneously. The purpose of this study was to test the transmission of the Atypical/Nor98 scrapie agent in three genotypes of Suffolk sheep and characterize the distribution of misfolded prion protein (PrPSc). Ten sheep were intracranially inoculated with brain homogenate from a sheep with Atypical/Nor98 scrapie. All sheep with the ARQ/ARQ and ARQ/ARR genotypes developed Atypical/Nor98 scrapie confirmed by immunohistochemistry, and one sheep with the VRQ/ARQ genotype had detectable PrPSc consistent with Atypical/Nor98 scrapie at the experimental endpoint of 8 years. Sheep with mild early accumulations of PrPSc in the cerebellum had concomitant retinal PrPSc. Accordingly, large amounts of retinal PrPSc were identified in clinically affected sheep and sheep with dense accumulations of PrPSc in the cerebellum.

Introduction

Atypical/Nor98 scrapie (AS) is a fatal prion disease of sheep caused by a misfolded form of the prion protein. Unlike classical scrapie (CS), AS is thought to be a spontaneously occurring disease [1–3]. This is supported by the presence of AS in countries that are free of classical scrapie [4, 5]. It typically affects a single older sheep within a flock, and cases of AS are sporadic and isolated suggesting that natural transmission is unlikely.

The susceptibility of sheep to CS is closely related to polymorphisms in the prion protein gene (PRNP) [6, 7]. Polymorphisms associated with susceptibility or resistance to CS occur at codons 136, 154, and 171. Sheep with the V136R154Q171 and A136R154Q171 haplotypes are susceptible to CS; however, the amino acid polymorphisms A136, R154, and R171 are associated with relative resistance [8–10]. Conversely, naturally occurring cases of AS arise in sheep with the AHQ, ARQ, and ARR haplotypes, and a polymorphism substituting phenylalanine (F) at codon 141 in the PRNP gene increases the risk of AS [11–13].

Several experiments have demonstrated the ability of the AS agent to transmit within the natural host after intracranial inoculation [14–16]. One study found that the AS agent could transmit after a high oral dose of AS brain homogenate [17]. Nonetheless, AS is still considered unlikely to transmit under field conditions; therefore, eradication and surveillance programs for CS have allowed exceptions for AS. As research into AS unfolds, the biological relevance of this disease is gaining attention. Two studies have demonstrated phenotype changes in AS that imply a possible origin for classical scrapie [18] and classical BSE [19]. The present study was designed to generate AS brain material for subsequent projects to investigate interspecies transmission events. Herein, we report our findings after the experimental transmission of AS in sheep with the VRQ/ARQ, ARQ/ARQ, and ARQ/ARR genotypes. This study validates previous work on these genotypes and documents the early accumulation of PrPSc in the retina of sheep with AS.

Results and discussion

All three genotypes of sheep, VRQ/ARQ, ARQ/ARQ, and ARQ/ARR, were susceptible to the AS agent after intracranial inoculation of donor brain homogenate. The diagnosis of AS was confirmed by enzyme immunoassay (EIA) and immunohistochemistry (IHC) with the latter being confirmative. Previous studies have demonstrated experimental transmission of AS to AHQ/AHQ [14, 15] and ARQ/ARQ [16] genotype sheep after intracerebral transmission. Another study showed a phenotypic shift from AS to CH1641-like classical scrapie in a sheep with the AHQ/AHQ genotype [18]. In this study, sheep with the ARQ/ARR genotype had the shortest incubation period ranging from 4.9 years to the experimental endpoint of 8 years (Table 1), and the attack rate was 100% (5/5). Clinical signs were observed in all ARQ/ARR sheep except for a single wether that was culled early to help establish experimental endpoints. Three ARQ/ARR genotype sheep were euthanized due to clinical neurologic disease 4.9–6.7 years post-inoculation. Out of the three genotypes examined, only the ARQ/ARR genotype sheep developed clinical neurologic disease within the eight-year incubation period. In clinically neurologic sheep, we observed stiff legged and hypermetric ataxia (dysmetria), abnormal rear stance, generalized tremors, tremors of the lips, weight loss, and generalized malaise. The spectrum of clinical signs was comparable to other reports of experimental AS in sheep [14, 15]. Three ARQ/ARR genotype sheep (804, 927 and 948) with the most severe dysmetria also had the greatest amount of cerebellar PrPSc. Since dysmetria is typical of animals with cerebellar disease [20], the tendency to observe this as the most consistent and severe neurologic sign is likely related to the characteristic cerebellar accumulation of PrPSc in sheep with AS. The ARQ/ARQ genotype had a long incubation period and remained clinically asymptomatic, as also reported by Okada et al. [16].

Table 1. Results of atypical scrapie transmission in Suffolk sheep. https://doi.org/10.1371/journal.pone.0246503.t001

Sheep with the ARQ/ARQ genotype were positive for AS PrPSc by IHC (2/2). One positive wether remained asymptomatic and was necropsied at the experimental endpoint; whereas, the other sheep was culled due to intercurrent disease around four years post-inoculation. Out of the three original VRQ/ARQ genotype sheep, a single presymptomatic wether had PrPSc in the cerebellum and retina at the experimental endpoint of 8.1 years. The other two sheep succumbed to intercurrent disease, and they did not have detectable PrPSc by means of IHC or EIA at 1.2- and 2.9-years post-inoculation. The VRQ allele, that is generally associated with susceptibility to classical scrapie, is usually absent from naturally occurring AS cases [5, 12, 21]. However, in a study of AS cases from Great Britain, a single VRQ/ARQ case was reported [22]. The prolonged incubation period after intracranial inoculation of AS in a VRQ/ARQ genotype sheep is compatible with the low prevalence in field cases of AS. In fact, field cases of AS often have a polymorphism substituting phenylalanine (F) at codon 141 in the PRNP gene, and most cases have either the AF141RQ or AHQ alleles [12]. All of the sheep in this study contained the amino acid leucine (L) at codon 141.

In order to confirm that sheep had AS and rule out concomitant infection with classical scrapie, all tissues were examined by IHC for PrPSc. The distribution of PrPSc in the brains of sheep was consistent with AS. Immunolabeling of PrPSc appeared as granular and punctate deposits and was largely restricted to the molecular layer of the cerebellum (Fig 1A). Small amounts of punctate and granular staining were also seen in the cerebral cortex, basal nuclei, thalamus, and midbrain. In classical scrapie, PrPSc is found in the dorsal motor nucleus of the vagus nerve (DMNV), one of the early sites of central nervous system accumulation, and in the lymphoid tissue [3]. In the present experiment, PrPSc was observed in the spinal trigeminal tract (Fig 1B), and there was a lack of staining for PrPSc in the DMNV (Fig 1C). Additionally, no PrPSc was detectable by IHC in the lymphoid or peripheral tissues of any sheep; it remained confined to the CNS. Other studies have demonstrated infectivity in peripheral and lymphoid tissues that were IHC negative [17, 23]. This distribution of PrPSc in the present study was consistent with AS in sheep [1, 14]. Furthermore, all genotypes of sheep had similar PrPSc distributions; however, the density of staining was less severe in asymptomatic ARQ/ARQ and VRQ/ARQ genotype sheep. Given a longer incubation period culminating in clinical disease, it is expected that these genotypes would develop more severe PrPSc deposition similar to ARQ/ARR genotype sheep. PrPSc was also found in the spinal cords of each genotype of sheep. Staining of PrPSc appeared as small particulate or fine granular deposits in the dorsal horn. In sheep with the ARQ/ARR genotype, there was minimal PrPSc and it was usually observed in the cervical cord alone. Sheep 929 that lived to the experimental endpoint had PrPSc in both the cervical and thoracic cord segments. In sheep 958 (ARQ/ARQ) and 943 (VRQ/ARQ), there was a mild amount of PrPSc in the dorsal horn of the cervical, thoracic, and lumbar spinal cord segments. This differs from classical scrapie that involves the entire grey matter of the spinal cord in late stage disease [24].

Fig 1. Immunoreactivity of PrPSc in sheep with atypical scrapie.

(A) There is a large amount of PrPSc (red color) within the molecular layer of cerebellum in sheep 958 (ARQ/ARQ). (B) PrPSc (red color) is confined to the spinal trigeminal tract in the medulla oblongata in sheep 948 (ARQ/ARR). (C) The dorsal motor nucleus of the vagus nerve (circle) is devoid of PrPSc in sheep 948. (D) There are multifocal patchy aggregates of PrPSc (red color) in the molecular layer of the cerebellum in sheep 943 (VRQ/ARQ). (E) A small amount of PrPSc (red color) is present in the retina of sheep 943. (F) In sheep 958 there are large amounts of PrPSc (red color) in the plexiform layers of the retina.


We performed both IHC and EIA on cerebellum, cerebrum (parietal cortex), and medulla oblongata at the level of the obex. For the ARQ/ARR and ARQ/ARQ genotype sheep, there was 100% (7/7) agreement between IHC and EIA at detecting PrPSc in the cerebellum. In contrast, the single positive VRQ/ARQ sheep was IHC positive and EIA negative in the cerebellum. This discrepancy was presumably due to the patchy and sparse distribution of PrPSc in the cerebellum (Fig 1D). PrPSc was rarely observed in other brain regions of this animal; however, PrPSc was detected in the retina with IHC (Fig 1E). Moreover, retinal PrPSc was present in each genotype of sheep with atypical scrapie PrPSc in the cerebellum. In clinical sheep with abundant cerebellar PrPSc, there were large amounts of PrPSc in the retina (Fig 1F). PrPSc occurred mostly in the inner and outer plexiform layers, but some minimal labeling was seen in the ganglion and nuclear cell layers. Other reports that describe atypical scrapie do not report retinal PrPSc [1–5, 14–16, 25, 26]. In this study, sheep intracranially inoculated with the atypical scrapie agent accumulated retinal PrPSc in the early stages of disease concomitant with cerebellar PrPSc. This is significant because, sequentially, retinal PrPSc accumulates early in disease; therefore, IHC of retinal tissue may be more sensitive compared to non-cerebellar brain regions.

This experiment demonstrated the transmission of atypical scrapie to three genotypes of sheep after intracranially inoculation, and it is the first study demonstrating experimental transmission to sheep with a VRQ/ARQ PRNP genotype. Additionally, atypical scrapie is further characterized by demonstrating early accumulation of PrPSc in the retina of experimentally inoculated sheep.

Materials and methods Animals for this experiment were derived from a known scrapie-free flock at the United States Department of Agriculture National Animal Disease Center in Ames, IA. This study used ten Suffolk sheep, nine wethers and one ewe. Nine sheep were 1 year old at the time of inoculation. A single sheep, #958, was 2 years old. Sheep in this study had three distinct PRNP genotypes: ARQ/ARQ, ARQ/ARR, and VRQ/ARQ. The genotypes were determined using polymerase chain reaction and Sanger sequencing as previously described [27]. Sheep were homozygous at other known polymorphic sites M112, G127, M137, S138, L141, R151, M157, N176, H180, Q189, T195, T196, R211, Q220, and R223.

The inoculum for this experiment was cerebral homogenate from an AHQ/ARH genotype sheep with atypical scrapie from Norway (Hedalen). The inoculum was obtained through a collaboration with Sylvie Benestad at the Norwegian Veterinary Institute. The brain homogenate was prepared as a 10% w/v homogenate. Sheep were intracranially inoculated with 1 ml (0.1 grams) of brain homogenate. The procedure has been described previously [28]. Briefly, the sheep were anesthetized with xylazine and a surgical field was prepped over the junction of parietal and frontal bones. A 1-cm skin incision was made, and then a 1-mm hole was drilled along the midline of the calvaria. A 9-cm spinal needle was inserted through the hole, and the inoculum was injected into the cranium. Sheep were kept in a biosecurity level 2 indoor pen for two weeks following inoculation and then moved to an outdoor area. They were fed a daily ration of pelleted and loose alfalfa hay. Sheep were monitored daily for any maladies or other clinical signs consistent with scrapie. The experimental endpoint for this experiment included the earliest of either unequivocal neurologic disease or 8 years post-inoculation. The final 8-year endpoint was established by performing a preliminarily cull of sheep 933 to help determine an appropriate endpoint. Sheep were euthanized at the onset of clinical disease or untreatable intercurrent disease. The method of euthanasia was intravenous administration of sodium pentobarbital as per label directions or as directed by an animal resources attending veterinarian. Clinical signs of disease included abnormalities in gate and/or stance, and ataxia.

A full post-mortem examination was performed on each sheep, and a routine set of tissues were collected consistent with previous experiments [29, 30]. A duplicate set of the following tissues were frozen or saved to 10% buffered neutral formalin: brain, spinal cord, pituitary, trigeminal ganglia, eyes, sciatic nerve, third eyelid, palatine tonsil, pharyngeal tonsil, lymph nodes (mesenteric, retropharyngeal, prescapular, and popliteal), spleen, esophagus, forestomaches, intestines, rectal mucosa, thymus, liver, kidney, urinary bladder, pancreas, salivary gland, thyroid gland, adrenal gland, trachea, lung, turbinate, nasal planum, heart, tongue, masseter, diaphragm, triceps brachii, biceps femoris, and psoas major. Formalin fixed tissues were processed, paraffin embedded, and sectioned at optimal thickness (brain, 4 μm; lymphoid, 3 μm; and other, 5 μm) for hematoxylin and eosin staining and IHC. For IHC, a cocktail of the monoclonal anti-PrPSc antibodies F89/160.1.5 [31] and F99/97.6.1 [32] was applied at a concentration of 5 μg/mL using an automated stainer. Frozen portions of cerebellum, parietal cerebral cortex, and brainstem at the level of the obex were homogenized and tested for the presence of PrPSc using a commercially available EIA (HerdChek; IDEXX Laboratories, Westbrook, ME) according to kit instructions.

Ethics statement

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Published: 31 August 2021

Classical BSE prions emerge from asymptomatic pigs challenged with atypical/Nor98 scrapie

Belén Marín, Alicia Otero, Séverine Lugan, Juan Carlos Espinosa, Alba Marín-Moreno, Enric Vidal, Carlos Hedman, Antonio Romero, Martí Pumarola, Juan J. Badiola, Juan María Torres, Olivier Andréoletti & Rosa Bolea 

Scientific Reports volume 11, Article number: 17428 (2021) Cite this article

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Abstract

Pigs are susceptible to infection with the classical bovine spongiform encephalopathy (C-BSE) agent following experimental inoculation, and PrPSc accumulation was detected in porcine tissues after the inoculation of certain scrapie and chronic wasting disease isolates. However, a robust transmission barrier has been described in this species and, although they were exposed to C-BSE agent in many European countries, no cases of natural transmissible spongiform encephalopathies (TSE) infections have been reported in pigs. Transmission of atypical scrapie to bovinized mice resulted in the emergence of C-BSE prions. Here, we conducted a study to determine if pigs are susceptible to atypical scrapie. To this end, 12, 8–9-month-old minipigs were intracerebrally inoculated with two atypical scrapie sources. Animals were euthanized between 22- and 72-months post inoculation without clinical signs of TSE. All pigs tested negative for PrPSc accumulation by enzyme immunoassay, immunohistochemistry, western blotting and bioassay in porcine PrP mice. Surprisingly, in vitro protein misfolding cyclic amplification demonstrated the presence of C-BSE prions in different brain areas from seven pigs inoculated with both atypical scrapie isolates. Our results suggest that pigs exposed to atypical scrapie prions could become a reservoir for C-BSE and corroborate that C-BSE prions emerge during interspecies passage of atypical scrapie.

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In conclusion, our findings suggest that, although pigs present a strong transmission barrier against the propagation of atypical scrapie, they can propagate low levels of C-BSE prions. The prevalence of atypical scrapie and the presence of infectivity in tissues from atypical scrapie infected sheep are underestimated24,25. Given that pigs have demonstrated being susceptible to other prion diseases, and to propagate prions without showing signs of disease, the measures implemented to ban the inclusion of ruminant proteins in livestock feed must not be interrupted.


Update on chronic wasting disease (CWD) III

EFSA Panel on Biological Hazards (BIOHAZ),Kostas Koutsoumanis,Ana Allende,Avelino Alvarez-Ordoňez,Declan Bolton,Sara Bover-Cid,Marianne Chemaly,Robert Davies,Alessandra De Cesare,Lieve Herman,Friederike Hilbert,Roland Lindqvist,Maarten Nauta,Luisa Peixe,Giuseppe Ru,Panagiotis Skandamis,Elisabetta Suffredini,Olivier Andreoletti,Sylvie L Benestad,Emmanuel Comoy,Romolo Nonno,Teresa da Silva Felicio,Angel Ortiz-Pelaez,Marion M Simmons, … 

First published: 11 November 2019 https://doi.org/10.2903/j.efsa.2019.5863

3.2.1.2 Non‐cervid domestic species

The remarkably high rate of natural CWD transmission in the ongoing NA epidemics raises the question of the risk to livestock grazing on CWD‐contaminated shared rangeland and subsequently developing a novel CWD‐related prion disease. This issue has been investigated by transmitting CWD via experimental challenge to cattle, sheep and pigs and to tg mouse lines expressing the relevant species PrP.

For cattle challenged with CWD, PrPSc was detected in approximately 40% of intracerebrally inoculated animals (Hamir et al., 2005, 2006a, 2007). Tg mice expressing bovine PrP have also been challenged with CWD and while published studies have negative outcomes (Tamguney et al., 2009b), unpublished data provided for the purposes of this Opinion indicate that some transmission of individual isolates to bovinised mice is possible (Table 1).

In small ruminant recipients, a low rate of transmission was reported between 35 and 72 months post‐infection (mpi) in ARQ/ARQ and ARQ/VRQ sheep intracerebrally challenged with mule deer CWD (Hamir et al., 2006b), while two out of two ARQ/ARQ sheep intracerebrally inoculated with elk CWD developed clinical disease after 28 mpi (Madsen‐Bouterse et al., 2016). However, tg mice expressing ARQ sheep PrP were resistant (Tamguney et al., 2006) and tg mice expressing the VRQ PrP allele were poorly susceptible to clinical disease (Beringue et al., 2012; Madsen‐Bouterse et al., 2016). In contrast, tg mice expressing VRQ sheep PrP challenged with CWD have resulted in highly efficient, life‐long asymptomatic replication of these prions in the spleen tissue (Beringue et al., 2012).

A recent study investigated the potential for swine to serve as hosts of the CWD agent(s) by intracerebral or oral challenge of crossbred piglets (Moore et al., 2016b, 2017). Pigs sacrificed at 6 mpi, approximately the age at which pigs reach market weight, were clinically healthy and negative by diagnostic tests, although low‐level CWD agent replication could be detected in the CNS by bioassay in tg cervinised mice. Among pigs that were incubated for up to 73 mpi, some gave diagnostic evidence of CWD replication in the brain between 42 and 72 mpi. Importantly, this was observed also in one orally challenged pig at 64 mpi and the presence of low‐level CWD replication was confirmed by mouse bioassay. The authors of this study argued that pigs can support low‐level amplification of CWD prions, although the species barrier to CWD infection is relatively high and that the detection of infectivity in orally inoculated pigs with a mouse bioassay raises the possibility that naturally exposed pigs could act as a reservoir of CWD infectivity.

Research Project: Pathobiology, Genetics, and Detection of Transmissible Spongiform Encephalopathies Location: Virus and Prion Research

Title: Experimental transmission of the chronic wasting disease agent to swine after oral or intracranial inoculation

Author item MOORE, SARAH - Orise Fellow item WEST GREENLEE, M - Iowa State University item KONDRU, NAVEEN - Iowa State University item MANNE, SIREESHA - Iowa State University item Smith, Jodi item Kunkle, Robert item KANTHASAMY, ANUMANTHA - Iowa State University item Greenlee, Justin Submitted to: Journal of Virology Publication Type: Peer Reviewed Journal Publication Acceptance Date: 7/6/2017 Publication Date: 9/12/2017

Citation: Moore, S.J., West Greenlee, M.H., Kondru, N., Manne, S., Smith, J.D., Kunkle, R.A., Kanthasamy, A., Greenlee, J.J. 2017. Experimental transmission of the chronic wasting disease agent to swine after oral or intracranial inoculation. Journal of Virology. 91(19):e00926-17. https://doi.org/10.1128/JVI.00926-17.

Interpretive Summary: Chronic wasting disease (CWD) is a fatal disease of wild and captive deer and elk that causes damaging changes in the brain. The infectious agent is an abnormal protein called a prion that has misfolded from its normal state. Whether CWD can transmit to swine is unknown. This study evaluated the potential of pigs to develop CWD after either intracranial or oral inoculation. Our data indicates that swine do accumulate the abnormal prion protein associated with CWD after intracranial or oral inoculation. Further, there was evidence of abnormal prion protein accumulation in lymph nodes. Currently, swine rations in the U.S. could contain animal derived components including materials from deer or elk. In addition, feral swine could be exposed to infected carcasses in areas where CWD is present in wildlife populations. This information is useful to wildlife managers and individuals in the swine and captive cervid industries. These findings could impact future regulations for the disposal of offal from deer and elk slaughtered in commercial operations. U.S. regulators should carefully consider the new information from this study before relaxing feed ban standards designed to control potentially feed borne prion diseases.

Technical Abstract: Chronic wasting disease (CWD) is a naturally occurring, fatal neurodegenerative disease of cervids. The potential for swine to serve as a host for the agent of chronic wasting disease is unknown. The purpose of this study was to investigate the susceptibility of swine to the CWD agent following oral or intracranial experimental inoculation. Crossbred piglets were assigned to one of three groups: intracranially inoculated (n=20), orally inoculated (n=19), or non-inoculated (n=9). At approximately the age at which commercial pigs reach market weight, half of the pigs in each group were culled ('market weight' groups). The remaining pigs ('aged' groups) were allowed to incubate for up to 73 months post inoculation (MPI). Tissues collected at necropsy were examined for disease-associated prion protein (PrPSc) by western blotting (WB), antigen-capture immunoassay (EIA), immunohistochemistry (IHC) and in vitro real-time quaking induced conversion (RT-QuIC). Brain samples from selected pigs were also bioassayed in mice expressing porcine prion protein. Four intracranially inoculated aged pigs and one orally inoculated aged pig were positive by EIA, IHC and/or WB. Using RT-QuIC, PrPSc was detected in lymphoid and/or brain tissue from pigs in all inoculated groups. Bioassay was positive in 4 out of 5 pigs assayed. This study demonstrates that pigs can serve as hosts for CWD, though with scant PrPSc accumulation requiring sensitive detection methods. Detection of infectivity in orally inoculated pigs using mouse bioassay raises the possibility that naturally exposed pigs could act as a reservoir of CWD infectivity.


12 September 2017

Experimental Transmission of the Chronic Wasting Disease Agent to Swine after Oral or Intracranial Inoculation

Authors: S. Jo Moore, M. Heather West Greenlee, Naveen Kondru, Sireesha Manne, Jodi D. Smith, Robert A. Kunkle, Anumantha Kanthasamy, and Justin J. Greenlee 


AUTHORS INFO & AFFILIATIONS


Volume 91, Number 19

1 October 2017

ABSTRACT

ABSTRACT

Chronic wasting disease (CWD) is a naturally occurring, fatal neurodegenerative disease of cervids. The potential for swine to serve as hosts for the agent of CWD is unknown. The purpose of this study was to investigate the susceptibility of swine to the CWD agent following experimental oral or intracranial inoculation. Crossbred piglets were assigned to three groups, intracranially inoculated (n = 20), orally inoculated (n = 19), and noninoculated (n = 9). At approximately the age at which commercial pigs reach market weight, half of the pigs in each group were culled (“market weight” groups). The remaining pigs (“aged” groups) were allowed to incubate for up to 73 months postinoculation (mpi). Tissues collected at necropsy were examined for disease-associated prion protein (PrPSc) by Western blotting (WB), antigen capture enzyme immunoassay (EIA), immunohistochemistry (IHC), and in vitro real-time quaking-induced conversion (RT-QuIC). Brain samples from selected pigs were also bioassayed in mice expressing porcine prion protein. Four intracranially inoculated aged pigs and one orally inoculated aged pig were positive by EIA, IHC, and/or WB. By RT-QuIC, PrPSc was detected in lymphoid and/or brain tissue from one or more pigs in each inoculated group. The bioassay was positive in four out of five pigs assayed. This study demonstrates that pigs can support low-level amplification of CWD prions, although the species barrier to CWD infection is relatively high. However, detection of infectivity in orally inoculated pigs with a mouse bioassay raises the possibility that naturally exposed pigs could act as a reservoir of CWD infectivity. IMPORTANCE We challenged domestic swine with the chronic wasting disease agent by inoculation directly into the brain (intracranially) or by oral gavage (orally). Disease-associated prion protein (PrPSc) was detected in brain and lymphoid tissues from intracranially and orally inoculated pigs as early as 8 months of age (6 months postinoculation). Only one pig developed clinical neurologic signs suggestive of prion disease. The amount of PrPSc in the brains and lymphoid tissues of positive pigs was small, especially in orally inoculated pigs. Regardless, positive results obtained with orally inoculated pigs suggest that it may be possible for swine to serve as a reservoir for prion disease under natural conditions.


Research Project: TRANSMISSION, DIFFERENTIATION, AND PATHOBIOLOGY OF TRANSMISSIBLE SPONGIFORM ENCEPHALOPATHIES Location: Virus and Prion Research

Title: Disease-associated prion protein detected in lymphoid tissues from pigs challenged with the agent of chronic wasting disease 

Conclusions: This study demonstrates that PrPSc accumulates in lymphoid tissues from pigs challenged intracranially or orally with the CWD agent, and can be detected as early as 4 months after challenge. CWD-infected pigs rarely develop clinical disease and if they do, they do so after a long incubation period. This raises the possibility that CWD-infected pigs could shed prions into their environment long before they develop clinical disease. Furthermore, lymphoid tissues from CWD-infected pigs could present a potential source of CWD infectivity in the animal and human food chains.



Friday, December 14, 2012 

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

snip..... 

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

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

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

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

The amount of animal PAP that is of deer and/or elk origin imported from the USA to GB can not be determined, however, as it is not specified in TRACES. 

It may constitute a small percentage of the 8412 kilos of non-fish origin processed animal proteins that were imported from US into GB in 2011. 

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

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

snip..... 

36% in 2007 (Almberg et al., 2011). In such areas, population declines of deer of up to 30 to 50% have been observed (Almberg et al., 2011). In areas of Colorado, the prevalence can be as high as 30% (EFSA, 2011). The clinical signs of CWD in affected adults are weight loss and behavioural changes that can span weeks or months (Williams, 2005). In addition, signs might include excessive salivation, behavioural alterations including a fixed stare and changes in interaction with other animals in the herd, and an altered stance (Williams, 2005). These signs are indistinguishable from cervids experimentally infected with bovine spongiform encephalopathy (BSE). Given this, if CWD was to be introduced into countries with BSE such as GB, for example, infected deer populations would need to be tested to differentiate if they were infected with CWD or BSE to minimise the risk of BSE entering the human food-chain via affected venison. snip..... The rate of transmission of CWD has been reported to be as high as 30% and can approach 100% among captive animals in endemic areas (Safar et al., 2008). 

snip..... 

In summary, in endemic areas, there is a medium probability that the soil and surrounding environment is contaminated with CWD prions and in a bioavailable form. In rural areas where CWD has not been reported and deer are present, there is a greater than negligible risk the soil is contaminated with CWD prion. snip..... In summary, given the volume of tourists, hunters and servicemen moving between GB and North America, the probability of at least one person travelling to/from a CWD affected area and, in doing so, contaminating their clothing, footwear and/or equipment prior to arriving in GB is greater than negligible... For deer hunters, specifically, the risk is likely to be greater given the increased contact with deer and their environment. However, there is significant uncertainty associated with these estimates. 

snip..... 

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

snip..... 


***> READ THIS VERY, VERY, CAREFULLY, AUGUST 1997 MAD COW FEED BAN WAS A SHAM, AS I HAVE STATED SINCE 1997! 3 FAILSAFES THE FDA ET AL PREACHED AS IF IT WERE THE GOSPEL, IN TERMS OF MAD COW BSE DISEASE IN USA, AND WHY IT IS/WAS/NOT A PROBLEM FOR THE USA, and those are; 

BSE TESTING (failed terribly and proven to be a sham) 

BSE SURVEILLANCE (failed terribly and proven to be a sham) 

BSE 589.2001 FEED REGULATIONS (another colossal failure, and proven to be a sham) 

these are facts folks. trump et al just admitted it with the feed ban. 

see; 

FDA Reports on VFD Compliance 

John Maday 

August 30, 2019 09:46 AM VFD-Form 007 (640x427) 

Before and after the current Veterinary Feed Directive rules took full effect in January, 2017, the FDA focused primarily on education and outreach. ( John Maday ) Before and after the current Veterinary Feed Directive (VFD) rules took full effect in January, 2017, the FDA focused primarily on education and outreach to help feed mills, veterinarians and producers understand and comply with the requirements. Since then, FDA has gradually increased the number of VFD inspections and initiated enforcement actions when necessary. On August 29, FDA released its first report on inspection and compliance activities. The report, titled “Summary Assessment of Veterinary Feed Directive Compliance Activities Conducted in Fiscal Years 2016 – 2018,” is available online.


SUNDAY, SEPTEMBER 1, 2019 

***> FDA Reports on VFD Compliance 


TUESDAY, APRIL 18, 2017 

*** EXTREME USA FDA PART 589 TSE PRION FEED LOOP HOLE STILL EXIST, AND PRICE OF POKER GOES UP *** 



2020

Monday, November 30, 2020 

Tunisia has become the second country after Algeria to detect a case of CPD Camel Prion Disease within a year 

REPORT OF THE MEETING OF THE OIE SCIENTIFIC COMMISSION FOR ANIMAL DISEASES Paris, 9–13 September 2019

Scientific Commission/September 2019

Tunisia has become the second country after Algeria to detect a case of CPD within a year

10.2. Prion disease in dromedary camels 


TUESDAY, SEPTEMBER 07, 2021 

Atypical Bovine Spongiform Encephalopathy BSE OIE, FDA 589.2001 FEED REGULATIONS, and Ingestion Therefrom


ZOONOSIS OF SCRAPIE TSE PRION

O.05: Transmission of prions to primates after extended silent incubation periods: Implications for BSE and scrapie risk assessment in human populations 

Emmanuel Comoy, Jacqueline Mikol, Valerie Durand, Sophie Luccantoni, Evelyne Correia, Nathalie Lescoutra, Capucine Dehen, and Jean-Philippe Deslys Atomic Energy Commission; Fontenay-aux-Roses, France 

Prion diseases (PD) are the unique neurodegenerative proteinopathies reputed to be transmissible under field conditions since decades. The transmission of Bovine Spongiform Encephalopathy (BSE) to humans evidenced that an animal PD might be zoonotic under appropriate conditions. Contrarily, in the absence of obvious (epidemiological or experimental) elements supporting a transmission or genetic predispositions, PD, like the other proteinopathies, are reputed to occur spontaneously (atpical animal prion strains, sporadic CJD summing 80% of human prion cases). 

Non-human primate models provided the first evidences supporting the transmissibiity of human prion strains and the zoonotic potential of BSE. Among them, cynomolgus macaques brought major information for BSE risk assessment for human health (Chen, 2014), according to their phylogenetic proximity to humans and extended lifetime. We used this model to assess the zoonotic potential of other animal PD from bovine, ovine and cervid origins even after very long silent incubation periods. 

*** We recently observed the direct transmission of a natural classical scrapie isolate to macaque after a 10-year silent incubation period, 

***with features similar to some reported for human cases of sporadic CJD, albeit requiring fourfold long incubation than BSE. Scrapie, as recently evoked in humanized mice (Cassard, 2014), 

***is the third potentially zoonotic PD (with BSE and L-type BSE), 

***thus questioning the origin of human sporadic cases. 

We will present an updated panorama of our different transmission studies and discuss the implications of such extended incubation periods on risk assessment of animal PD for human health. 

=============== 

***thus questioning the origin of human sporadic cases*** 

=============== 

***our findings suggest that possible transmission risk of H-type BSE to sheep and human. Bioassay will be required to determine whether the PMCA products are infectious to these animals. 

============== 


***Transmission data also revealed that several scrapie prions propagate in HuPrP-Tg mice with efficiency comparable to that of cattle BSE. While the efficiency of transmission at primary passage was low, subsequent passages resulted in a highly virulent prion disease in both Met129 and Val129 mice. 

***Transmission of the different scrapie isolates in these mice leads to the emergence of prion strain phenotypes that showed similar characteristics to those displayed by MM1 or VV2 sCJD prion. 

***These results demonstrate that scrapie prions have a zoonotic potential and raise new questions about the possible link between animal and human prions. 

 
PRION 2016 TOKYO

Saturday, April 23, 2016

SCRAPIE WS-01: Prion diseases in animals and zoonotic potential 2016

Prion. 10:S15-S21. 2016 ISSN: 1933-6896 printl 1933-690X online

Taylor & Francis

Prion 2016 Animal Prion Disease Workshop Abstracts

WS-01: Prion diseases in animals and zoonotic potential

Juan Maria Torres a, Olivier Andreoletti b, J uan-Carlos Espinosa a. Vincent Beringue c. Patricia Aguilar a,

Natalia Fernandez-Borges a. and Alba Marin-Moreno a

"Centro de Investigacion en Sanidad Animal ( CISA-INIA ). Valdeolmos, Madrid. Spain; b UMR INRA -ENVT 1225 Interactions Holes Agents Pathogenes. ENVT. Toulouse. France: "UR892. Virologie lmmunologie MolécuIaires, Jouy-en-Josas. France

Dietary exposure to bovine spongiform encephalopathy (BSE) contaminated bovine tissues is considered as the origin of variant Creutzfeldt Jakob (vCJD) disease in human. To date, BSE agent is the only recognized zoonotic prion... Despite the variety of Transmissible Spongiform Encephalopathy (TSE) agents that have been circulating for centuries in farmed ruminants there is no apparent epidemiological link between exposure to ruminant products and the occurrence of other form of TSE in human like sporadic Creutzfeldt Jakob Disease (sCJD). However, the zoonotic potential of the diversity of circulating TSE agents has never been systematically assessed. The major issue in experimental assessment of TSEs zoonotic potential lies in the modeling of the ‘species barrier‘, the biological phenomenon that limits TSE agents’ propagation from a species to another. In the last decade, mice genetically engineered to express normal forms of the human prion protein has proved essential in studying human prions pathogenesis and modeling the capacity of TSEs to cross the human species barrier.

To assess the zoonotic potential of prions circulating in farmed ruminants, we study their transmission ability in transgenic mice expressing human PrPC (HuPrP-Tg). Two lines of mice expressing different forms of the human PrPC (129Met or 129Val) are used to determine the role of the Met129Val dimorphism in susceptibility/resistance to the different agents.

These transmission experiments confirm the ability of BSE prions to propagate in 129M- HuPrP-Tg mice and demonstrate that Met129 homozygotes may be susceptible to BSE in sheep or goat to a greater degree than the BSE agent in cattle and that these agents can convey molecular properties and neuropathological indistinguishable from vCJD. However homozygous 129V mice are resistant to all tested BSE derived prions independently of the originating species suggesting a higher transmission barrier for 129V-PrP variant.

Transmission data also revealed that several scrapie prions propagate in HuPrP-Tg mice with efficiency comparable to that of cattle BSE. While the efficiency of transmission at primary passage was low, subsequent passages resulted in a highly virulent prion disease in both Met129 and Val129 mice. 

Transmission of the different scrapie isolates in these mice leads to the emergence of prion strain phenotypes that showed similar characteristics to those displayed by MM1 or VV2 sCJD prion. 

These results demonstrate that scrapie prions have a zoonotic potential and raise new questions about the possible link between animal and human prions. 

 
***> why do we not want to do TSE transmission studies on chimpanzees $

5. A positive result from a chimpanzee challenged severly would likely create alarm in some circles even if the result could not be interpreted for man. 

***> I have a view that all these agents could be transmitted provided a large enough dose by appropriate routes was given and the animals kept long enough. 

***> Until the mechanisms of the species barrier are more clearly understood it might be best to retain that hypothesis.

snip...

R. BRADLEY


Title: Transmission of scrapie prions to primate after an extended silent incubation period) 

*** In complement to the recent demonstration that humanized mice are susceptible to scrapie, we report here the first observation of direct transmission of a natural classical scrapie isolate to a macaque after a 10-year incubation period. Neuropathologic examination revealed all of the features of a prion disease: spongiform change, neuronal loss, and accumulation of PrPres throughout the CNS. 

*** This observation strengthens the questioning of the harmlessness of scrapie to humans, at a time when protective measures for human and animal health are being dismantled and reduced as c-BSE is considered controlled and being eradicated. 

*** Our results underscore the importance of precautionary and protective measures and the necessity for long-term experimental transmission studies to assess the zoonotic potential of other animal prion strains. 


***> Moreover, sporadic disease has never been observed in breeding colonies or primate research laboratories, most notably among hundreds of animals over several decades of study at the National Institutes of Health25, and in nearly twenty older animals continuously housed in our own facility. <***

Transmission of scrapie prions to primate after an extended silent incubation period 

Emmanuel E. Comoy, Jacqueline Mikol, Sophie Luccantoni-Freire, Evelyne Correia, Nathalie Lescoutra-Etchegaray, Valérie Durand, Capucine Dehen, Olivier Andreoletti, Cristina Casalone, Juergen A. Richt, Justin J. Greenlee, Thierry Baron, Sylvie L. Benestad, Paul Brown & Jean-Philippe Deslys Scientific Reports volume 5, Article number: 11573 (2015) | Download Citation

Abstract 

Classical bovine spongiform encephalopathy (c-BSE) is the only animal prion disease reputed to be zoonotic, causing variant Creutzfeldt-Jakob disease (vCJD) in humans and having guided protective measures for animal and human health against animal prion diseases. Recently, partial transmissions to humanized mice showed that the zoonotic potential of scrapie might be similar to c-BSE. We here report the direct transmission of a natural classical scrapie isolate to cynomolgus macaque, a highly relevant model for human prion diseases, after a 10-year silent incubation period, with features similar to those reported for human cases of sporadic CJD. Scrapie is thus actually transmissible to primates with incubation periods compatible with their life expectancy, although fourfold longer than BSE. Long-term experimental transmission studies are necessary to better assess the zoonotic potential of other prion diseases with high prevalence, notably Chronic Wasting Disease of deer and elk and atypical/Nor98 scrapie.

SNIP...

Discussion We describe the transmission of spongiform encephalopathy in a non-human primate inoculated 10 years earlier with a strain of sheep c-scrapie. Because of this extended incubation period in a facility in which other prion diseases are under study, we are obliged to consider two alternative possibilities that might explain its occurrence. We first considered the possibility of a sporadic origin (like CJD in humans). Such an event is extremely improbable because the inoculated animal was 14 years old when the clinical signs appeared, i.e. about 40% through the expected natural lifetime of this species, compared to a peak age incidence of 60–65 years in human sporadic CJD, or about 80% through their expected lifetimes. Moreover, sporadic disease has never been observed in breeding colonies or primate research laboratories, most notably among hundreds of animals over several decades of study at the National Institutes of Health25, and in nearly twenty older animals continuously housed in our own facility.

The second possibility is a laboratory cross-contamination. Three facts make this possibility equally unlikely. First, handling of specimens in our laboratory is performed with fastidious attention to the avoidance of any such cross-contamination. Second, no laboratory cross-contamination has ever been documented in other primate laboratories, including the NIH, even between infected and uninfected animals housed in the same or adjacent cages with daily intimate contact (P. Brown, personal communication). Third, the cerebral lesion profile is different from all the other prion diseases we have studied in this model19, with a correlation between cerebellar lesions (massive spongiform change of Purkinje cells, intense PrPres staining and reactive gliosis26) and ataxia. The iron deposits present in the globus pallidus are a non specific finding that have been reported previously in neurodegenerative diseases and aging27. Conversely, the thalamic lesion was reminiscent of a metabolic disease due to thiamine deficiency28 but blood thiamine levels were within normal limits (data not shown). The preferential distribution of spongiform change in cortex associated with a limited distribution in the brainstem is reminiscent of the lesion profile in MM2c and VV1 sCJD patients29, but interspecies comparison of lesion profiles should be interpreted with caution. It is of note that the same classical scrapie isolate induced TSE in C57Bl/6 mice with similar incubation periods and lesional profiles as a sample derived from a MM1 sCJD patient30.

We are therefore confident that the illness in this cynomolgus macaque represents a true transmission of a sheep c-scrapie isolate directly to an old-world monkey, which taxonomically resides in the primate subdivision (parvorder of catarrhini) that includes humans. With an homology of its PrP protein with humans of 96.4%31, cynomolgus macaque constitutes a highly relevant model for assessing zoonotic risk of prion diseases. Since our initial aim was to show the absence of transmission of scrapie to macaques in the worst-case scenario, we obtained materials from a flock of naturally-infected sheep, affecting animals with different genotypes32. This c-scrapie isolate exhibited complete transmission in ARQ/ARQ sheep (332 ± 56 days) and Tg338 transgenic mice expressing ovine VRQ/VRQ prion protein (220 ± 5 days) (O. Andreoletti, personal communication). From the standpoint of zoonotic risk, it is important to note that sheep with c-scrapie (including the isolate used in our study) have demonstrable infectivity throughout their lymphoreticular system early in the incubation period of the disease (3 months-old for all the lymphoid organs, and as early as 2 months-old in gut-associated lymph nodes)33. In addition, scrapie infectivity has been identified in blood34, milk35 and skeletal muscle36 from asymptomatic but scrapie infected small ruminants which implies a potential dietary exposure for consumers.

Two earlier studies have reported the occurrence of clinical TSE in cynomolgus macaques after exposures to scrapie isolates. In the first study, the “Compton” scrapie isolate (derived from an English sheep) and serially propagated for 9 passages in goats did not transmit TSE in cynomolgus macaque, rhesus macaque or chimpanzee within 7 years following intracerebral challenge1; conversely, after 8 supplementary passages in conventional mice, this “Compton” isolate induced TSE in a cynomolgus macaque 5 years after intracerebral challenge, but rhesus macaques and chimpanzee remained asymptomatic 8.5 years post-exposure8. However, multiple successive passages that are classically used to select laboratory-adapted prion strains can significantly modify the initial properties of a scrapie isolate, thus questioning the relevance of zoonotic potential for the initial sheep-derived isolate. The same isolate had also induced disease into squirrel monkeys (new-world monkey)9. A second historical observation reported that a cynomolgus macaque developed TSE 6 years post-inoculation with brain homogenate from a scrapie-infected Suffolk ewe (derived from USA), whereas a rhesus macaque and a chimpanzee exposed to the same inoculum remained healthy 9 years post-exposure1. This inoculum also induced TSE in squirrel monkeys after 4 passages in mice. Other scrapie transmission attempts in macaque failed but had more shorter periods of observation in comparison to the current study. Further, it is possible that there are differences in the zoonotic potential of different scrapie strains.

The most striking observation in our study is the extended incubation period of scrapie in the macaque model, which has several implications. Firstly, our observations constitute experimental evidence in favor of the zoonotic potential of c-scrapie, at least for this isolate that has been extensively studied32,33,34,35,36. The cross-species zoonotic ability of this isolate should be confirmed by performing duplicate intracerebral exposures and assessing the transmissibility by the oral route (a successful transmission of prion strains through the intracerebral route may not necessarily indicate the potential for oral transmission37). However, such confirmatory experiments may require more than one decade, which is hardly compatible with current general management and support of scientific projects; thus this study should be rather considered as a case report.

Secondly, transmission of c-BSE to primates occurred within 8 years post exposure for the lowest doses able to transmit the disease (the survival period after inoculation is inversely proportional to the initial amount of infectious inoculum). The occurrence of scrapie 10 years after exposure to a high dose (25 mg) of scrapie-infected sheep brain suggests that the macaque has a higher species barrier for sheep c-scrapie than c-BSE, although it is notable that previous studies based on in vitro conversion of PrP suggested that BSE and scrapie prions would have a similar conversion potential for human PrP38.

Thirdly, prion diseases typically have longer incubation periods after oral exposure than after intracerebral inoculations: since humans can develop Kuru 47 years after oral exposure39, an incubation time of several decades after oral exposure to scrapie would therefore be expected, leading the disease to occur in older adults, i.e. the peak age for cases considered to be sporadic disease, and making a distinction between scrapie-associated and truly sporadic disease extremely difficult to appreciate.

Fourthly, epidemiologic evidence is necessary to confirm the zoonotic potential of an animal disease suggested by experimental studies. A relatively short incubation period and a peculiar epidemiological situation (e.g., all the first vCJD cases occurring in the country with the most important ongoing c-BSE epizootic) led to a high degree of suspicion that c-BSE was the cause of vCJD. Sporadic CJD are considered spontaneous diseases with an almost stable and constant worldwide prevalence (0.5–2 cases per million inhabitants per year), and previous epidemiological studies were unable to draw a link between sCJD and classical scrapie6,7,40,41, even though external causes were hypothesized to explain the occurrence of some sCJD clusters42,43,44. However, extended incubation periods exceeding several decades would impair the predictive values of epidemiological surveillance for prion diseases, already weakened by a limited prevalence of prion diseases and the multiplicity of isolates gathered under the phenotypes of “scrapie” and “sporadic CJD”.

Fifthly, considering this 10 year-long incubation period, together with both laboratory and epidemiological evidence of decade or longer intervals between infection and clinical onset of disease, no premature conclusions should be drawn from negative transmission studies in cynomolgus macaques with less than a decade of observation, as in the aforementioned historical transmission studies of scrapie to primates1,8,9. Our observations and those of others45,46 to date are unable to provide definitive evidence regarding the zoonotic potential of CWD, atypical/Nor98 scrapie or H-type BSE. The extended incubation period of the scrapie-affected macaque in the current study also underscores the limitations of rodent models expressing human PrP for assessing the zoonotic potential of some prion diseases since their lifespan remains limited to approximately two years21,47,48. This point is illustrated by the fact that the recently reported transmission of scrapie to humanized mice was not associated with clinical signs for up to 750 days and occurred in an extreme minority of mice with only a marginal increase in attack rate upon second passage13. The low attack rate in these studies is certainly linked to the limited lifespan of mice compared to the very long periods of observation necessary to demonstrate the development of scrapie. Alternatively, one could estimate that a successful second passage is the result of strain adaptation to the species barrier, thus poorly relevant of the real zoonotic potential of the original scrapie isolate of sheep origin49. The development of scrapie in this primate after an incubation period compatible with its lifespan complements the study conducted in transgenic (humanized) mice; taken together these studies suggest that some isolates of sheep scrapie can promote misfolding of the human prion protein and that scrapie can develop within the lifespan of some primate species.

In addition to previous studies on scrapie transmission to primate1,8,9 and the recently published study on transgenic humanized mice13, our results constitute new evidence for recommending that the potential risk of scrapie for human health should not be dismissed. Indeed, human PrP transgenic mice and primates are the most relevant models for investigating the human transmission barrier. To what extent such models are informative for measuring the zoonotic potential of an animal TSE under field exposure conditions is unknown. During the past decades, many protective measures have been successfully implemented to protect cattle from the spread of c-BSE, and some of these measures have been extended to sheep and goats to protect from scrapie according to the principle of precaution. Since cases of c-BSE have greatly reduced in number, those protective measures are currently being challenged and relaxed in the absence of other known zoonotic animal prion disease. We recommend that risk managers should be aware of the long term potential risk to human health of at least certain scrapie isolates, notably for lymphotropic strains like the classical scrapie strain used in the current study. Relatively high amounts of infectivity in peripheral lymphoid organs in animals infected with these strains could lead to contamination of food products produced for human consumption. Efforts should also be maintained to further assess the zoonotic potential of other animal prion strains in long-term studies, notably lymphotropic strains with high prevalence like CWD, which is spreading across North America, and atypical/Nor98 scrapie (Nor98)50 that was first detected in the past two decades and now represents approximately half of all reported cases of prion diseases in small ruminants worldwide, including territories previously considered as scrapie free... Even if the prevailing view is that sporadic CJD is due to the spontaneous formation of CJD prions, it remains possible that its apparent sporadic nature may, at least in part, result from our limited capacity to identify an environmental origin.


1: J Infect Dis 1980 Aug;142(2):205-8

Oral transmission of kuru, Creutzfeldt-Jakob disease, and scrapie to nonhuman primates.

Gibbs CJ Jr, Amyx HL, Bacote A, Masters CL, Gajdusek DC.

Kuru and Creutzfeldt-Jakob disease of humans and scrapie disease of sheep and goats were transmitted to squirrel monkeys (Saimiri sciureus) that were exposed to the infectious agents only by their nonforced consumption of known infectious tissues. The asymptomatic incubation period in the one monkey exposed to the virus of kuru was 36 months; that in the two monkeys exposed to the virus of Creutzfeldt-Jakob disease was 23 and 27 months, respectively; and that in the two monkeys exposed to the virus of scrapie was 25 and 32 months, respectively. Careful physical examination of the buccal cavities of all of the monkeys failed to reveal signs or oral lesions. One additional monkey similarly exposed to kuru has remained asymptomatic during the 39 months that it has been under observation.

snip...

The successful transmission of kuru, Creutzfeldt-Jakob disease, and scrapie by natural feeding to squirrel monkeys that we have reported provides further grounds for concern that scrapie-infected meat may occasionally give rise in humans to Creutzfeldt-Jakob disease.

PMID: 6997404


Recently the question has again been brought up as to whether scrapie is transmissible to man. This has followed reports that the disease has been transmitted to primates. One particularly lurid speculation (Gajdusek 1977) conjectures that the agents of scrapie, kuru, Creutzfeldt-Jakob disease and transmissible encephalopathy of mink are varieties of a single "virus". The U.S. Department of Agriculture concluded that it could "no longer justify or permit scrapie-blood line and scrapie-exposed sheep and goats to be processed for human or animal food at slaughter or rendering plants" (ARC 84/77)" The problem is emphasised by the finding that some strains of scrapie produce lesions identical to the once which characterise the human dementias"

Whether true or not. the hypothesis that these agents might be transmissible to man raises two considerations. First, the safety of laboratory personnel requires prompt attention. Second, action such as the "scorched meat" policy of USDA makes the solution of the acrapie problem urgent if the sheep industry is not to suffer grievously.

snip...

76/10.12/4.6


Nature. 1972 Mar 10;236(5341):73-4.

Transmission of scrapie to the cynomolgus monkey (Macaca fascicularis).

Gibbs CJ Jr, Gajdusek DC.

Nature 236, 73 - 74 (10 March 1972); doi:10.1038/236073a0

Transmission of Scrapie to the Cynomolgus Monkey (Macaca fascicularis)

C. J. GIBBS jun. & D. C. GAJDUSEK

National Institute of Neurological Diseases and Stroke, National Institutes of Health, Bethesda, Maryland

SCRAPIE has been transmitted to the cynomolgus, or crab-eating, monkey (Macaca fascicularis) with an incubation period of more than 5 yr from the time of intracerebral inoculation of scrapie-infected mouse brain. The animal developed a chronic central nervous system degeneration, with ataxia, tremor and myoclonus with associated severe scrapie-like pathology of intensive astroglial hypertrophy and proliferation, neuronal vacuolation and status spongiosus of grey matter. The strain of scrapie virus used was the eighth passage in Swiss mice (NIH) of a Compton strain of scrapie obtained as ninth intracerebral passage of the agent in goat brain, from Dr R. L. Chandler (ARC, Compton, Berkshire).


Norway Regulatory process VKM order for CWD TSE Prion after discovery on the Hardangervidda in 2020

Regulatory process VKM order for CWD after discovery on the Hardangervidda in 2020 

Published 09.10.2020 Last changed 09.10.2020 Print

Scrapie (CWD) has been detected in a wild reindeer buck on the Hardangervidda. The diagnosis was made on 10.9.2020. The disease has previously been detected in the Nordfjella wild reindeer area, and VKM has prepared several reports in that connection.

In the wake of the new detection on the Hardangervidda, the Norwegian Food Safety Authority and the Norwegian Environment Agency need an assessment regarding updated knowledge about the disease and risk factors for the spread of the disease inside and out of the wild reindeer area etc.

Since it is urgent to get answers to some questions, we have chosen to divide the assignment into two phases. Attached is the mandate for the order of phase 1 on scrapie from the Norwegian Environment Agency and the Norwegian Food Safety Authority. Key words for questions in phase 2 are food safety, game management, etc. We will return to this.

Timeline for this work: Consultation of VKM order Before the final order is sent, we want input on the draft order for a new assessment

Consultation deadline: 05.10.2020

https://www.mattilsynet.no/dyr_og_dyrehold/dyrehelse/dyresykdommer/skrantesjuke__cwd_/vkmbestilling_om_cwd_etter_funn_paa_hardangervidda_i_2020.40531?utm_campaign=Nyhetsbrev&utm_medium=Epost&utm_source=Mattilsynet&utm_term=VKMbestilling_om_CWD_etter_funn_paa_Hardangervidda_i_2020&utm_content=Dyrehelse




FRIDAY, OCTOBER 09, 2020 

Norway Regulatory process VKM order for CWD TSE Prion after discovery on the Hardangervidda in 2020


MONDAY, SEPTEMBER 14, 2020 

Norway Chronic Wasting Disease (CWD) identified in a wild reindeer at Hardanger Plateau


FRIDAY, SEPTEMBER 11, 2020 

Norway Skrantesjuke CWD TSE Prion detected on reindeer buck from Hardangervidda


WEDNESDAY, APRIL 01, 2020 

Norway Chronic Wasting Disease CWD TSE Prion Skrantesjuke 2 Positive Moose for 2019


MONDAY, NOVEMBER 18, 2019 

Norway Chronic Wasting Disease CWD TSE Prion Detected in Sixth Moose


WEDNESDAY, MARCH 06, 2019 

Norway The Madness Continues in Nordfjella Chronic Wasting Disease CWD TSE Prion


THURSDAY, FEBRUARY 14, 2019 

Norway Eradication of Chronic Wasting Disease is not completed 


THURSDAY, SEPTEMBER 26, 2019
Sweden The third case of CWD in moose in Arjeplog is now established https://chronic-wasting-disease.blogspot.com/2019/09/sweden-third-case-of-cwd-in-moose-in.html
SATURDAY, JUNE 01, 2019 

Sweden Documents Another Case of Chronic Wasting Disease CWD TSE Prion Norrbotten


FRIDAY, APRIL 12, 2019 

Sweden Wasting Disease (CWD) discovered on moose in Norrbotten County

SATURDAY, MARCH 10, 2018 FINLAND REPORTS FIRST CASE OF CHRONIC WASTING DISEASE CWD TSE PRION IN A moose or European elk (Alces alces) http://chronic-wasting-disease.blogspot.com/2018/03/finland-reports-first-case-of-chronic.html

TUESDAY, FEBRUARY 09, 2021 

Sweden Confirms First Detection of Chronic Wasting Disease in Moose (Alces alces)


SATURDAY, JANUARY 9, 2021 

A case-control study of scrapie Nor98 in Norwegian sheep flocks


TUESDAY, SEPTEMBER 21, 2021 

Detection of CWD prions in naturally infected white-tailed deer fetuses and gestational tissues by PMCA 


TUESDAY, AUGUST 31, 2021 

***> TEXAS CHRONIC WASTING DISEASE CWD TSE PRION HAS CONFIRMED 260 CASES TO DATE <***


***> 1st and foremost your biggest problem is 'VOLUNTARY'! AS with the BSE 589.2001 FEED REGULATIONS, especially since it is still voluntary with cervid, knowing full well that cwd and scrapie will transmit to pigs by oral route. VOLUNTARY DOES NOT WORK! all animal products should be banned and be made mandatory, and the herd certification program should be mandatory, or you don't move cervid. IF THE CWD HERD CERTIFICATION IS NOT MANDATORY, it will be another colossal tse prion failure from the start.

***> 2nd USA should declare a Declaration of Extraordinary Emergency due to CWD, and all exports of cervid and cervid products must be stopped internationally, and there should be a ban of interstate movement of cervid, until a live cwd test is available.

***> 3rd Captive Farmed cervid ESCAPEES should be made mandatory to report immediately, and strict regulations for those suspect cwd deer that just happen to disappear. IF a cervid escapes and is not found, that farm should be indefinitely shut down, all movement, until aid MIA cervid is found, and if not ever found, that farm shut down permanently.

***> 4th Captive Farmed Cervid, INDEMNITY, NO MORE Federal indemnity program, or what i call, ENTITLEMENT PROGRAM for game farm industry. NO MORE BAIL OUTS FROM TAX PAYERS. if the captive industry can't buy insurance to protect not only themselves, but also their customers, and especially the STATE, from Chronic Wasting Disease CWD TSE Prion or what some call mad deer disease and harm therefrom, IF they can't afford to buy that insurance that will cover all of it, then they DO NOT GET A PERMIT to have a game farm for anything. This CWD TSE Prion can/could/has caused property values to fall from some reports in some places. roll the dice, how much is a state willing to lose?

***> 5th QUARANTINE OF ALL FARMED CAPTIVE, BREEDERS, URINE, ANTLER, VELVET, SPERM, OR ANY FACILITY, AND THEIR PRODUCTS, that has been confirmed to have Chronic Wasting Disease CWD TSE Prion, the QUARANTINE should be for 21 years due to science showing what scrapie can do. 5 years is NOT near long enough. see; Infectious agent of sheep scrapie may persist in the environment for at least 16 to 21 years.

***> 6th America BSE 589.2001 FEED REGULATIONS CWD TSE Prion

***> 7TH TRUCKING TRANSPORTING CERVID CHRONIC WASTING DISEASE TSE PRION VIOLATING THE LACEY ACT

***> 8TH ALL CAPTIVE FARMING CERVID OPERATIONS MUST BE INSURED TO PAY FOR ANY CLEAN UP OF CWD AND QUARANTINE THERE FROM FOR THE STATE, NO MORE ENTITLEMENT PROGRAM FOR CERVID GAME FARMING PAY TO PLAY FOR CWD TSE PRION OFF THE TAX PAYERS BACK.

***> 9TH ANY STATE WITH DOCUMENTED CWD, INTERSTATE, NATIONAL, AND INTERNATIONAL MOVEMENT OF ALL CERVID, AND ALL CERVID PRODUCTS MUST BE HALTED!

***> 10TH BAN THE SALE OF STRAW BRED BUCKS AND ALL CERVID SEMEN AND URINE PRODUCTS

***> 11th ALL CAPTIVE FARMED CERVID AND THEIR PRODUCTS MUST BE CWD TSE PRION TESTED ANNUALLY AND BEFORE SALE FOR CWD TSE PRION

SEE FULL SCIENCE REFERENCES AND REASONINGS ;

Control of Chronic Wasting Disease OMB Control Number: 0579-0189 APHIS-2021-0004 Singeltary Submission



Docket No. APHIS-2018-0011 Chronic Wasting Disease Herd Certification



5 or 6 years quarantine is NOT LONG ENOUGH FOR CWD TSE PRION !!!

QUARANTINE NEEDS TO BE 21 YEARS FOR CWD TSE PRION !

FRIDAY, APRIL 30, 2021 

Should Property Evaluations Contain Scrapie, CWD, TSE PRION Environmental Contamination of the land?

***> Confidential!!!!

***> As early as 1992-3 there had been long studies conducted on small pastures containing scrapie infected sheep at the sheep research station associated with the Neuropathogenesis Unit in Edinburgh, Scotland. Whether these are documented...I don't know. But personal recounts both heard and recorded in a daily journal indicate that leaving the pastures free and replacing the topsoil completely at least 2 feet of thickness each year for SEVEN years....and then when very clean (proven scrapie free) sheep were placed on these small pastures.... the new sheep also broke out with scrapie and passed it to offspring. I am not sure that TSE contaminated ground could ever be free of the agent!! A very frightening revelation!!!

---end personal email---end...tss


WEDNESDAY, DECEMBER 04, 2013 

Chronic Wasting Disease CWD and Land Value concerns? 


***> This is very likely to have parallels with control efforts for CWD in cervids.

Paper

Rapid recontamination of a farm building occurs after attempted prion removal

Kevin Christopher Gough BSc (Hons), PhD Claire Alison Baker BSc (Hons) Steve Hawkins MIBiol Hugh Simmons BVSc, MRCVS, MBA, MA Timm Konold DrMedVet, PhD, MRCVS … See all authors 

First published: 19 January 2019 https://doi.org/10.1136/vr.105054

Abstract

The transmissible spongiform encephalopathy scrapie of sheep/goats and chronic wasting disease of cervids are associated with environmental reservoirs of infectivity. Preventing environmental prions acting as a source of infectivity to healthy animals is of major concern to farms that have had outbreaks of scrapie and also to the health management of wild and farmed cervids. Here, an efficient scrapie decontamination protocol was applied to a farm with high levels of environmental contamination with the scrapie agent. Post‐decontamination, no prion material was detected within samples taken from the farm buildings as determined using a sensitive in vitro replication assay (sPMCA). A bioassay consisting of 25 newborn lambs of highly susceptible prion protein genotype VRQ/VRQ introduced into this decontaminated barn was carried out in addition to sampling and analysis of dust samples that were collected during the bioassay. Twenty‐four of the animals examined by immunohistochemical analysis of lymphatic tissues were scrapie‐positive during the bioassay, samples of dust collected within the barn were positive by month 3. The data illustrates the difficulty in decontaminating farm buildings from scrapie, and demonstrates the likely contribution of farm dust to the recontamination of these environments to levels that are capable of causing disease.

snip...

This study clearly demonstrates the difficulty in removing scrapie infectivity from the farm environment. Practical and effective prion decontamination methods are still urgently required for decontamination of scrapie infectivity from farms that have had cases of scrapie and this is particularly relevant for scrapiepositive goatherds, which currently have limited genetic resistance to scrapie within commercial breeds.24 This is very likely to have parallels with control efforts for CWD in cervids.


***>This is very likely to have parallels with control efforts for CWD in cervids.







***> Infectious agent of sheep scrapie may persist in the environment for at least 16 years

***> Nine of these recurrences occurred 14–21 years after culling, apparently as the result of environmental contamination, but outside entry could not always be absolutely excluded. 

JOURNAL OF GENERAL VIROLOGY Volume 87, Issue 12

Infectious agent of sheep scrapie may persist in the environment for at least 16 years Free

Gudmundur Georgsson1, Sigurdur Sigurdarson2, Paul Brown3


Saturday, January 5, 2019 

Rapid recontamination of a farm building occurs after attempted prion removal 


The effectiveness of on-farm decontamination methods for scrapie - SE1865

Description

Scrapie infectivity persists on farms where infected animals have been removed1. Recently we have demonstrated that it is possible to detect environmental scrapie contamination biochemically using serial Protein Misfolding Cyclic Amplification (sPMCA)2, allowing the monitoring of scrapie infectivity on farm premises. Ongoing Defra study SE1863 has compared pen decontamination regimes on a scrapie-infected farm by both sheep bioassay and sPMCA. For bioassay, scrapie-free genetically susceptible lambs were introduced into pens decontaminated using distinct methodologies, all pens contained scrapie-positive lambs within 1 year. Remarkably this included lambs housed within a pen which had been jet washed/chloros treated, followed by regalvanisation/ replacement of all metalwork and painting of all other surfaces.

We have recently demonstrated using sPMCA, that material collected on swabs from vertical surfaces at heights inaccessible to sheep within a barn on the same scrapie affected farm contained scrapie prions (unpublished observations). We hypothesise that scrapie prions are most likely to have been deposited in these areas by bioaerosol movement. We propose that this bioaerosol movement contributes to scrapie transmission within the barn, and could account for the sheep that became positive within the pen containing re-galvanised/new metalwork and repainted surfaces (project SE1863). It is proposed that a thorough decontamination that would minimise prion-contaminated dust, both within the building and its immediate vicinity, is likely to increase the effectiveness of current methods for decontaminating farm buildings following outbreaks of scrapie. The proposed study builds on our previous data and will thoroughly investigate the potential for farm building scrapie-contamination via the bioaerosol route, a previously unrecognised route for dissemination of scrapie infectivity. This route could lead to the direct infection of healthy animals and/or indirect transmission of disease via contamination of surfaces within animal pens. The proposed study would analyse material collected using air samplers set up within “scrapie-infected” barns and their immediate vicinity, to confirm that prion containing material can be airborne within a scrapie infected farm environment. The study would incorporate a biochemical assessment of different surface decontamination methods, in order to demonstrate the best methodology and then the analysis of air and surface samples after a complete building decontamination to remove sources of dust and surface bound prions from both the building and its immediate vicinity. Analysis of such surface and air samples collected before and after treatment would measure the reduction in levels of infectivity. It is envisaged that the biochemical demonstration of airborne prions and the effective reduction in such prion dissemination would lead to a sheep bioassay experiment that would be conducted after a full farm decontamination. This would fully assess the effectiveness of an optimised scrapie decontamination strategy.

This study will contribute directly to Defra policy on best practice for on-farm decontamination after outbreaks of scrapie; a situation particularly relevant to decontamination after scrapie cases on goat farms where no genetic resistance to scrapie has currently been identified, and where complete decontamination is essential in order to stop recurrence of scrapie after restocking.

Objective

Phase 1

• Determine the presence and relative levels of airborne prions on a scrapie infected farm.

• Evaluate different pen surface decontamination procedures.

Phase 2

• Determine the presence of any airborne prions in a barn after a full decontamination.

Phase 3

• Further assess the efficacy of the decontamination procedure investigated in phase 2 by sheep bioassay.

Time-Scale and Cost

From: 2012 

To: 2016 

Cost: £326,784

Contractor / Funded Organisations

A D A S UK Ltd (ADAS)

Keywords Animals Fields of Study Animal Health


The Effectiveness of on-Farm Decontamination Methods for Scrapie

Institutions ADAS

Start date 2012

End date 2016

Objective Phase 1

Determine the presence and relative levels of airborne prions on a scrapie infected farm. Evaluate different pen surface decontamination procedures.

Phase 2

Determine the presence of any airborne prions in a barn after a full decontamination.

Phase 3

Further assess the efficacy of the decontamination procedure investigated in phase 2 by sheep bioassay.

More information

Scrapie infectivity persists on farms where infected animals have been removed1. Recently we have demonstrated that it is possible to detect environmental scrapie contamination biochemically using serial Protein Misfolding Cyclic Amplification (sPMCA)2, allowing the monitoring of scrapie infectivity on farm premises. Ongoing Defra study SE1863 has compared pen decontamination regimes on a scrapie-infected farm by both sheep bioassay and sPMCA. For bioassay, scrapie-free genetically susceptible lambs were introduced into pens decontaminated using distinct methodologies, all pens contained scrapie-positive lambs within 1 year. Remarkably this included lambs housed within a pen which had been jet washed/chloros treated, followed by regalvanisation/replacement of all metalwork and painting of all other surfaces.

We have recently demonstrated using sPMCA, that material collected on swabs from vertical surfaces at heights inaccessible to sheep within a barn on the same scrapie affected farm contained scrapie prions (unpublished observations). We hypothesise that scrapie prions are most likely to have been deposited in these areas by bioaerosol movement. We propose that this bioaerosol movement contributes to scrapie transmission within the barn, and could account for the sheep that became positive within the pen containing re-galvanised/new metalwork and repainted surfaces (project SE1863). It is proposed that a thorough decontamination that would minimise prion-contaminated dust, both within the building and its immediate vicinity, is likely to increase the effectiveness of current methods for decontaminating farm buildings following outbreaks of scrapie. The proposed study builds on our previous data and will thoroughly investigate the potential for farm building scrapie contamination via the bioaerosol route, a previously unrecognised route for dissemination of scrapie infectivity. This route could lead to the direct infection of healthy animals and/or indirect transmission of disease via contamination of surfaces within animal pens. The proposed study would analyse material collected using air samplers set up within “scrapie-infected” barns and their immediate vicinity, to confirm that prion containing material can be airborne within a scrapie infected farm environment. The study would incorporate a biochemical assessment of different surface decontamination methods, in order to demonstrate the best methodology and then the analysis of air and surface samples after a complete building decontamination to remove sources of dust and surface bound prions from both the building and its immediate vicinity. Analysis of such surface and air samples collected before and after treatment would measure the reduction in levels of infectivity. It is envisaged that the biochemical demonstration of airborne prions and the effective reduction in such prion dissemination would lead to a sheep bioassay experiment that would be conducted after a full farm decontamination. This would fully assess the effectiveness of an optimised scrapie decontamination strategy.

This study will contribute directly to Defra policy on best practice for on-farm decontamination after outbreaks of scrapie; a situation particularly relevant to decontamination after scrapie cases on goat farms where no genetic resistance to scrapie has currently been identified, and where complete decontamination is essential in order to stop recurrence of scrapie after restocking.

Funding Source

Department for Environment, Food and Rural Affairs

Project source

View this project

Project number

SE1865

Categories

Foodborne Disease

Policy and Planning 


Circulation of prions within dust on a scrapie affected farm

Kevin C Gough1 , Claire A Baker2 , Hugh A Simmons3 , Steve A Hawkins3 and Ben C Maddison2*

Abstract

Prion diseases are fatal neurological disorders that affect humans and animals. Scrapie of sheep/goats and Chronic Wasting Disease (CWD) of deer/elk are contagious prion diseases where environmental reservoirs have a direct link to the transmission of disease. Using protein misfolding cyclic amplification we demonstrate that scrapie PrPSc can be detected within circulating dusts that are present on a farm that is naturally contaminated with sheep scrapie. The presence of infectious scrapie within airborne dusts may represent a possible route of infection and illustrates the difficulties that may be associated with the effective decontamination of such scrapie affected premises.

snip... 

Discussion We present biochemical data illustrating the airborne movement of scrapie containing material within a contaminated farm environment. We were able to detect scrapie PrPSc within extracts from dusts collected over a 70 day period, in the absence of any sheep activity. We were also able to detect scrapie PrPSc within dusts collected within pasture at 30 m but not at 60 m distance away from the scrapie contaminated buildings, suggesting that the chance of contamination of pasture by scrapie contaminated dusts decreases with distance from contaminated farm buildings. PrPSc amplification by sPMCA has been shown to correlate with infectivity and amplified products have been shown to be infectious [14,15]. These experiments illustrate the potential for low dose scrapie infectivity to be present within such samples. We estimate low ng levels of scrapie positive brain equivalent were deposited per m2 over 70 days, in a barn previously occupied by sheep affected with scrapie. This movement of dusts and the accumulation of low levels of scrapie infectivity within this environment may in part explain previous observations where despite stringent pen decontamination regimens healthy lambs still became scrapie infected after apparent exposure from their environment alone [16]. The presence of sPMCA seeding activity and by inference, infectious prions within dusts, and their potential for airborne dissemination is highly novel and may have implications for the spread of scrapie within infected premises. The low level circulation and accumulation of scrapie prion containing dust material within the farm environment will likely impede the efficient decontamination of such scrapie contaminated buildings unless all possible reservoirs of dust are removed. Scrapie containing dusts could possibly infect animals during feeding and drinking, and respiratory and conjunctival routes may also be involved. It has been demonstrated that scrapie can be efficiently transmitted via the nasal route in sheep [17], as is also the case for CWD in both murine models and in white tailed deer [18-20].

The sources of dust borne prions are unknown but it seems reasonable to assume that faecal, urine, skin, parturient material and saliva-derived prions may contribute to this mobile environmental reservoir of infectivity. This work highlights a possible transmission route for scrapie within the farm environment, and this is likely to be paralleled in CWD which shows strong similarities with scrapie in terms of prion dissemination and disease transmission. The data indicate that the presence of scrapie prions in dust is likely to make the control of these diseases a considerable challenge.


Research Project: TRANSMISSION, DIFFERENTIATION, AND PATHOBIOLOGY OF TRANSMISSIBLE SPONGIFORM ENCEPHALOPATHIES Location: Virus and Prion Research

Title: Scrapie transmits to white-tailed deer by the oral route and has a molecular profile similar to chronic wasting disease

Author 

 item Greenlee, Justin item Moore, S - Orise Fellow item Smith, Jodi - Iowa State University item Kunkle, Robert item West Greenlee, M - Iowa State University Submitted to: American College of Veterinary Pathologists Meeting Publication Type: Abstract Only Publication Acceptance Date: 8/12/2015 Publication Date: N/A Citation: N/A

Interpretive Summary:

Technical Abstract: The purpose of this work was to determine susceptibility of white-tailed deer (WTD) to the agent of sheep scrapie and to compare the resultant PrPSc to that of the original inoculum and chronic wasting disease (CWD). We inoculated WTD by a natural route of exposure (concurrent oral and intranasal (IN); n=5) with a US scrapie isolate. All scrapie-inoculated deer had evidence of PrPSc accumulation. PrPSc was detected in lymphoid tissues at preclinical time points, and deer necropsied after 28 months post-inoculation had clinical signs, spongiform encephalopathy, and widespread distribution of PrPSc in neural and lymphoid tissues. Western blotting (WB) revealed PrPSc with 2 distinct molecular profiles. WB on cerebral cortex had a profile similar to the original scrapie inoculum, whereas WB of brainstem, cerebellum, or lymph nodes revealed PrPSc with a higher profile resembling CWD. Homogenates with the 2 distinct profiles from WTD with clinical scrapie were further passaged to mice expressing cervid prion protein and intranasally to sheep and WTD. In cervidized mice, the two inocula have distinct incubation times. Sheep inoculated intranasally with WTD derived scrapie developed disease, but only after inoculation with the inoculum that had a scrapie-like profile. The WTD study is ongoing, but deer in both inoculation groups are positive for PrPSc by rectal mucosal biopsy. In summary, this work demonstrates that WTD are susceptible to the agent of scrapie, two distinct molecular profiles of PrPSc are present in the tissues of affected deer, and inoculum of either profile readily passes to deer.


THE tse prion aka mad cow type disease is not your normal pathogen. 

The TSE prion disease survives ashing to 600 degrees celsius, that’s around 1112 degrees farenheit. 

you cannot cook the TSE prion disease out of meat. 

you can take the ash and mix it with saline and inject that ash into a mouse, and the mouse will go down with TSE. 

Prion Infected Meat-and-Bone Meal Is Still Infectious after Biodiesel Production as well. 

the TSE prion agent also survives Simulated Wastewater Treatment Processes. 

IN fact, you should also know that the TSE Prion agent will survive in the environment for years, if not decades. 

you can bury it and it will not go away. 

The TSE agent is capable of infected your water table i.e. Detection of protease-resistant cervid prion protein in water from a CWD-endemic area. 

it’s not your ordinary pathogen you can just cook it out and be done with. 

***> that’s what’s so worrisome about Iatrogenic mode of transmission, a simple autoclave will not kill this TSE prion agent.

1: J Neurol Neurosurg Psychiatry 1994 Jun;57(6):757-8 

***> Transmission of Creutzfeldt-Jakob disease to a chimpanzee by electrodes contaminated during neurosurgery. 

Gibbs CJ Jr, Asher DM, Kobrine A, Amyx HL, Sulima MP, Gajdusek DC. 

Laboratory of Central Nervous System Studies, National Institute of 

Neurological Disorders and Stroke, National Institutes of Health, 

Bethesda, MD 20892. 

Stereotactic multicontact electrodes used to probe the cerebral cortex of a middle aged woman with progressive dementia were previously implicated in the accidental transmission of Creutzfeldt-Jakob disease (CJD) to two younger patients. The diagnoses of CJD have been confirmed for all three cases. More than two years after their last use in humans, after three cleanings and repeated sterilisation in ethanol and formaldehyde vapour, the electrodes were implanted in the cortex of a chimpanzee. Eighteen months later the animal became ill with CJD. This finding serves to re-emphasise the potential danger posed by reuse of instruments contaminated with the agents of spongiform encephalopathies, even after scrupulous attempts to clean them. 

PMID: 8006664 [PubMed - indexed for MEDLINE] 


New studies on the heat resistance of hamster-adapted scrapie agent: Threshold survival after ashing at 600°C suggests an inorganic template of replication 


Prion Infected Meat-and-Bone Meal Is Still Infectious after Biodiesel Production 


MONDAY, APRIL 19, 2021

Evaluation of the application for new alternative biodiesel production process for rendered fat including Category 1 animal by-products (BDI-RepCat® process, AT) ???


Detection of protease-resistant cervid prion protein in water from a CWD-endemic area 


A Quantitative Assessment of the Amount of Prion Diverted to Category 1 Materials and Wastewater During Processing 


Rapid assessment of bovine spongiform encephalopathy prion inactivation by heat treatment in yellow grease produced in the industrial manufacturing process of meat and bone meals 


THURSDAY, FEBRUARY 28, 2019 

BSE infectivity survives burial for five years with only limited spread



***> CONGRESSIONAL ABSTRACTS PRION CONFERENCE 2018

P69 Experimental transmission of CWD from white-tailed deer to co-housed reindeer 

Mitchell G (1), Walther I (1), Staskevicius A (1), Soutyrine A (1), Balachandran A (1) 

(1) National & OIE Reference Laboratory for Scrapie and CWD, Canadian Food Inspection Agency, Ottawa, Ontario, Canada. 

Chronic wasting disease (CWD) continues to be detected in wild and farmed cervid populations of North America, affecting predominantly white-tailed deer, mule deer and elk. Extensive herds of wild caribou exist in northern regions of Canada, although surveillance has not detected the presence of CWD in this population. Oral experimental transmission has demonstrated that reindeer, a species closely related to caribou, are susceptible to CWD. Recently, CWD was detected for the first time in Europe, in wild Norwegian reindeer, advancing the possibility that caribou in North America could also become infected. Given the potential overlap in habitat between wild CWD-infected cervids and wild caribou herds in Canada, we sought to investigate the horizontal transmissibility of CWD from white-tailed deer to reindeer. 

Two white-tailed deer were orally inoculated with a brain homogenate prepared from a farmed Canadian white-tailed deer previously diagnosed with CWD. Two reindeer, with no history of exposure to CWD, were housed in the same enclosure as the white-tailed deer, 3.5 months after the deer were orally inoculated. The white-tailed deer developed clinical signs consistent with CWD beginning at 15.2 and 21 months post-inoculation (mpi), and were euthanized at 18.7 and 23.1 mpi, respectively. Confirmatory testing by immunohistochemistry (IHC) and western blot demonstrated widespread aggregates of pathological prion protein (PrPCWD) in the central nervous system and lymphoid tissues of both inoculated white-tailed deer. Both reindeer were subjected to recto-anal mucosal associated lymphoid tissue (RAMALT) biopsy at 20 months post-exposure (mpe) to the white-tailed deer. The biopsy from one reindeer contained PrPCWD confirmed by IHC. This reindeer displayed only subtle clinical evidence of disease prior to a rapid decline in condition requiring euthanasia at 22.5 mpe. Analysis of tissues from this reindeer by IHC revealed widespread PrPCWD deposition, predominantly in central nervous system and lymphoreticular tissues. Western blot molecular profiles were similar between both orally inoculated white-tailed deer and the CWD positive reindeer. Despite sharing the same enclosure, the other reindeer was RAMALT negative at 20 mpe, and PrPCWD was not detected in brainstem and lymphoid tissues following necropsy at 35 mpe. Sequencing of the prion protein gene from both reindeer revealed differences at several codons, which may have influenced susceptibility to infection. 

Natural transmission of CWD occurs relatively efficiently amongst cervids, supporting the expanding geographic distribution of disease and the potential for transmission to previously naive populations. The efficient horizontal transmission of CWD from white-tailed deer to reindeer observed here highlights the potential for reindeer to become infected if exposed to other cervids or environments infected with CWD. 

SOURCE REFERENCE 2018 PRION CONFERENCE ABSTRACT

Research Project: TRANSMISSION, DIFFERENTIATION, AND PATHOBIOLOGY OF TRANSMISSIBLE SPONGIFORM ENCEPHALOPATHIES Location: Virus and Prion Research

Title: Horizontal transmission of chronic wasting disease in reindeer

Author

item MOORE, SARAH - ORISE FELLOW item KUNKLE, ROBERT item WEST GREENLEE, MARY - IOWA STATE UNIVERSITY item Nicholson, Eric item RICHT, JUERGEN item HAMIR, AMIRALI item WATERS, WADE item Greenlee, Justin

Submitted to: Emerging Infectious Diseases

Publication Type: Peer Reviewed Journal

Publication Acceptance Date: 8/29/2016

Publication Date: 12/1/2016

Citation: Moore, S., Kunkle, R., Greenlee, M., Nicholson, E., Richt, J., Hamir, A., Waters, W., Greenlee, J. 2016. Horizontal transmission of chronic wasting disease in reindeer. Emerging Infectious Diseases. 22(12):2142-2145. doi:10.3201/eid2212.160635.

Interpretive Summary: Chronic wasting disease (CWD) is a fatal neurodegenerative disease that occurs in farmed and wild cervids (deer and elk) of North America and was recently diagnosed in a single free-ranging reindeer (Rangifer tarandus tarandus) in Norway. CWD is a transmissible spongiform encephalopathy (TSE) that is caused by infectious proteins called prions that are resistant to various methods of decontamination and environmental degradation. Little is known about the susceptibility of or potential for transmission amongst reindeer. In this experiment, we tested the susceptibility of reindeer to CWD from various sources (elk, mule deer, or white-tailed deer) after intracranial inoculation and tested the potential for infected reindeer to transmit to non-inoculated animals by co-housing or housing in adjacent pens. Reindeer were susceptible to CWD from elk, mule deer, or white-tailed deer sources after experimental inoculation. Most importantly, non-inoculated reindeer that were co-housed with infected reindeer or housed in pens adjacent to infected reindeer but without the potential for nose-to-nose contact also developed evidence of CWD infection. This is a major new finding that may have a great impact on the recently diagnosed case of CWD in the only remaining free-ranging reindeer population in Europe as our findings imply that horizontal transmission to other reindeer within that herd has already occurred. Further, this information will help regulatory and wildlife officials developing plans to reduce or eliminate CWD and cervid farmers that want to ensure that their herd remains CWD-free, but were previously unsure of the potential for reindeer to transmit CWD.

Technical Abstract: Chronic wasting disease (CWD) is a naturally-occurring, fatal prion disease of cervids. Reindeer (Rangifer tarandus tarandus) are susceptible to CWD following oral challenge, and CWD was recently reported in a free-ranging reindeer of Norway. Potential contact between CWD-affected cervids and Rangifer species that are free-ranging or co-housed on farms presents a potential risk of CWD transmission. The aims of this study were to 1) investigate the transmission of CWD from white-tailed deer (Odocoileus virginianus; CWDwtd), mule deer (Odocoileus hemionus; CWDmd), or elk (Cervus elaphus nelsoni; CWDelk) to reindeer via the intracranial route, and 2) to assess for direct and indirect horizontal transmission to non-inoculated sentinels. Three groups of 5 reindeer fawns were challenged intracranially with CWDwtd, CWDmd, or CWDelk. Two years after challenge of inoculated reindeer, non-inoculated negative control reindeer were introduced into the same pen as the CWDwtd inoculated reindeer (direct contact; n=4) or into a pen adjacent to the CWDmd inoculated reindeer (indirect contact; n=2). Experimentally inoculated reindeer were allowed to develop clinical disease. At death/euthanasia a complete necropsy examination was performed, including immunohistochemical testing of tissues for disease-associated CWD prion protein (PrPcwd). Intracranially challenged reindeer developed clinical disease from 21 months post-inoculation (months PI). PrPcwd was detected in 5 out of 6 sentinel reindeer although only 2 out of 6 developed clinical disease during the study period (< 57 months PI). We have shown that reindeer are susceptible to CWD from various cervid sources and can transmit CWD to naïve reindeer both directly and indirectly.


Infectivity surviving ashing to 600*C is (in my opinion) degradable but infective. based on Bown & Gajdusek, (1991), landfill and burial may be assumed to have a reduction factor of 98% (i.e. a factor of 50) over 3 years. CJD-infected brain-tissue remained infectious after storing at room-temperature for 22 months (Tateishi et al, 1988). Scrapie agent is known to remain viable after at least 30 months of desiccation (Wilson et al, 1950). and pastures that had been grazed by scrapie-infected sheep still appeared to be contaminated with scrapie agent three years after they were last occupied by sheep (Palsson, 1979).


Dr. Paul Brown Scrapie Soil Test BSE Inquiry Document


Using in vitro Prion replication for high sensitive detection of prions and prionlike proteins and for understanding mechanisms of transmission. 

Claudio Soto Mitchell Center for Alzheimer's diseases and related Brain disorders, Department of Neurology, University of Texas Medical School at Houston. 

Prion and prion-like proteins are misfolded protein aggregates with the ability to selfpropagate to spread disease between cells, organs and in some cases across individuals. I n T r a n s m i s s i b l e s p o n g i f o r m encephalopathies (TSEs), prions are mostly composed by a misfolded form of the prion protein (PrPSc), which propagates by transmitting its misfolding to the normal prion protein (PrPC). The availability of a procedure to replicate prions in the laboratory may be important to study the mechanism of prion and prion-like spreading and to develop high sensitive detection of small quantities of misfolded proteins in biological fluids, tissues and environmental samples. Protein Misfolding Cyclic Amplification (PMCA) is a simple, fast and efficient methodology to mimic prion replication in the test tube. PMCA is a platform technology that may enable amplification of any prion-like misfolded protein aggregating through a seeding/nucleation process. In TSEs, PMCA is able to detect the equivalent of one single molecule of infectious PrPSc and propagate prions that maintain high infectivity, strain properties and species specificity. Using PMCA we have been able to detect PrPSc in blood and urine of experimentally infected animals and humans affected by vCJD with high sensitivity and specificity. Recently, we have expanded the principles of PMCA to amplify amyloid-beta (Aβ) and alphasynuclein (α-syn) aggregates implicated in Alzheimer's and Parkinson's diseases, respectively. Experiments are ongoing to study the utility of this technology to detect Aβ and α-syn aggregates in samples of CSF and blood from patients affected by these diseases.

=========================

***>>> Recently, we have been using PMCA to study the role of environmental prion contamination on the horizontal spreading of TSEs. These experiments have focused on the study of the interaction of prions with plants and environmentally relevant surfaces. Our results show that plants (both leaves and roots) bind tightly to prions present in brain extracts and excreta (urine and feces) and retain even small quantities of PrPSc for long periods of time. Strikingly, ingestion of prioncontaminated leaves and roots produced disease with a 100% attack rate and an incubation period not substantially longer than feeding animals directly with scrapie brain homogenate. Furthermore, plants can uptake prions from contaminated soil and transport them to different parts of the plant tissue (stem and leaves). Similarly, prions bind tightly to a variety of environmentally relevant surfaces, including stones, wood, metals, plastic, glass, cement, etc. Prion contaminated surfaces efficiently transmit prion disease when these materials were directly injected into the brain of animals and strikingly when the contaminated surfaces were just placed in the animal cage. These findings demonstrate that environmental materials can efficiently bind infectious prions and act as carriers of infectivity, suggesting that they may play an important role in the horizontal transmission of the disease.

========================

Since its invention 13 years ago, PMCA has helped to answer fundamental questions of prion propagation and has broad applications in research areas including the food industry, blood bank safety and human and veterinary disease diagnosis. 

source reference Prion Conference 2015 abstract book

Grass Plants Bind, Retain, Uptake, and Transport Infectious Prions

Sandra Pritzkow,1 Rodrigo Morales,1 Fabio Moda,1,3 Uffaf Khan,1 Glenn C. Telling,2 Edward Hoover,2 and Claudio Soto1, * 1Mitchell Center for Alzheimer’s Disease and Related Brain Disorders, Department of Neurology, University of Texas Medical School at Houston, 6431 Fannin Street, Houston, TX 77030, USA

2Prion Research Center, Department of Microbiology, Immunology, and Pathology, College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, CO 80523, USA

3Present address: IRCCS Foundation Carlo Besta Neurological Institute, 20133 Milan, Italy *Correspondence: claudio.soto@uth.tmc.edu http://dx.doi.org/10.1016/j.celrep.2015.04.036

SUMMARY

Prions are the protein-based infectious agents responsible for prion diseases. Environmental prion contamination has been implicated in disease transmission. Here, we analyzed the binding and retention of infectious prion protein (PrPSc) to plants. Small quantities of PrPSc contained in diluted brain homogenate or in excretory materials (urine and feces) can bind to wheat grass roots and leaves. Wild-type hamsters were efficiently infected by ingestion of prion-contaminated plants. The prion-plant interaction occurs with prions from diverse origins, including chronic wasting disease. Furthermore, leaves contaminated by spraying with a prion-containing preparation retained PrPSc for several weeks in the living plant. Finally, plants can uptake prions from contaminated soil and transport them to aerial parts of the plant (stem and leaves). These findings demonstrate that plants can efficiently bind infectious prions and act as carriers of infectivity, suggesting a possible role of environmental prion contamination in the horizontal transmission of the disease.

INTRODUCTION

snip...

DISCUSSION

This study shows that plants can efficiently bind prions contained in brain extracts from diverse prion infected animals, including CWD-affected cervids. PrPSc attached to leaves and roots from wheat grass plants remains capable of seeding prion replication in vitro. Surprisingly, the small quantity of PrPSc naturally excreted in urine and feces from sick hamster or cervids was enough to efficiently contaminate plant tissue. Indeed, our results suggest that the majority of excreted PrPSc is efficiently captured by plants’ leaves and roots. Moreover, leaves can be contaminated by spraying them with a prion-containing extract, and PrPSc remains detectable in living plants for as long as the study was performed (several weeks). Remarkably, prion contaminated plants transmit prion disease to animals upon ingestion, producing a 100% attack rate and incubation periods not substantially longer than direct oral administration of sick brain homogenates.

Finally, an unexpected but exciting result was that plants were able to uptake prions from contaminated soil and transport them to aerial parts of the plant tissue. Although it may seem farfetched that plants can uptake proteins from the soil and transport it to the parts above the ground, there are already published reports of this phenomenon (McLaren et al., 1960; Jensen and McLaren, 1960;Paungfoo-Lonhienne et al., 2008). The high resistance of prions to degradation and their ability to efficiently cross biological barriers may play a role in this process. The mechanism by which plants bind, retain, uptake, and transport prions is unknown. We are currently studying the way in which prions interact with plants using purified, radioactively labeled PrPSc to determine specificity of the interaction, association constant, reversibility, saturation, movement, etc.

Epidemiological studies have shown numerous instances of scrapie or CWD recurrence upon reintroduction of animals on pastures previously exposed to prion-infected animals. Indeed, reappearance of scrapie has been documented following fallow periods of up to 16 years (Georgsson et al., 2006), and pastures were shown to retain infectious CWD prions for at least 2 years after exposure (Miller et al., 2004). It is likely that the environmentally mediated transmission of prion diseases depends upon the interaction of prions with diverse elements, including soil, water, environmental surfaces, various invertebrate animals, and plants.

However, since plants are such an important component of the environment and also a major source of food for many animal species, including humans, our results may have far-reaching implications for animal and human health. Currently, the perception of the riskfor animal-to-human prion transmission has beenmostly limited to consumption or exposure to contaminated meat; our results indicate that plants might also be an important vector of transmission that needs to be considered in risk assessment. 


RIGINAL RESEARCH ARTICLE

Front. Vet. Sci., 14 September 2015 | https://doi.org/10.3389/fvets.2015.00032

Objects in contact with classical scrapie sheep act as a reservoir for scrapie transmission

imageTimm Konold1*, imageStephen A. C. Hawkins2, imageLisa C. Thurston3, imageBen C. Maddison4, imageKevin C. Gough5, imageAnthony Duarte1 and imageHugh A. Simmons1

1Animal Sciences Unit, Animal and Plant Health Agency Weybridge, Addlestone, UK

2Pathology Department, Animal and Plant Health Agency Weybridge, Addlestone, UK

3Surveillance and Laboratory Services, Animal and Plant Health Agency Penrith, Penrith, UK

4ADAS UK, School of Veterinary Medicine and Science, University of Nottingham, Sutton Bonington, UK

5School of Veterinary Medicine and Science, University of Nottingham, Sutton Bonington, UK

Classical scrapie is an environmentally transmissible prion disease of sheep and goats. Prions can persist and remain potentially infectious in the environment for many years and thus pose a risk of infecting animals after re-stocking. In vitro studies using serial protein misfolding cyclic amplification (sPMCA) have suggested that objects on a scrapie-affected sheep farm could contribute to disease transmission. This in vivo study aimed to determine the role of field furniture (water troughs, feeding troughs, fencing, and other objects that sheep may rub against) used by a scrapie-infected sheep flock as a vector for disease transmission to scrapie-free lambs with the prion protein genotype VRQ/VRQ, which is associated with high susceptibility to classical scrapie. When the field furniture was placed in clean accommodation, sheep became infected when exposed to either a water trough (four out of five) or to objects used for rubbing (four out of seven). This field furniture had been used by the scrapie-infected flock 8 weeks earlier and had previously been shown to harbor scrapie prions by sPMCA. Sheep also became infected (20 out of 23) through exposure to contaminated field furniture placed within pasture not used by scrapie-infected sheep for 40 months, even though swabs from this furniture tested negative by PMCA. This infection rate decreased (1 out of 12) on the same paddock after replacement with clean field furniture. Twelve grazing sheep exposed to field furniture not in contact with scrapie-infected sheep for 18 months remained scrapie free. The findings of this study highlight the role of field furniture used by scrapie-infected sheep to act as a reservoir for disease re-introduction although infectivity declines considerably if the field furniture has not been in contact with scrapie-infected sheep for several months. PMCA may not be as sensitive as VRQ/VRQ sheep to test for environmental contamination.

snip...

Discussion 

Classical scrapie is an environmentally transmissible disease because it has been reported in naïve, supposedly previously unexposed sheep placed in pastures formerly occupied by scrapie-infected sheep (4, 19, 20). 

Although the vector for disease transmission is not known, soil is likely to be an important reservoir for prions (2) where – based on studies in rodents – prions can adhere to minerals as a biologically active form (21) and remain infectious for more than 2 years (22). 

Similarly, chronic wasting disease (CWD) has re-occurred in mule deer housed in paddocks used by infected deer 2 years earlier, which was assumed to be through foraging and soil consumption (23). 

Our study suggested that the risk of acquiring scrapie infection was greater through exposure to contaminated wooden, plastic, and metal surfaces via water or food troughs, fencing, and hurdles than through grazing. 

Drinking from a water trough used by the scrapie flock was sufficient to cause infection in sheep in a clean building. 

Exposure to fences and other objects used for rubbing also led to infection, which supported the hypothesis that skin may be a vector for disease transmission (9). 

The risk of these objects to cause infection was further demonstrated when 87% of 23 sheep presented with PrPSc in lymphoid tissue after grazing on one of the paddocks, which contained metal hurdles, a metal lamb creep and a water trough in contact with the scrapie flock up to 8 weeks earlier, whereas no infection had been demonstrated previously in sheep grazing on this paddock, when equipped with new fencing and field furniture. 

When the contaminated furniture and fencing were removed, the infection rate dropped significantly to 8% of 12 sheep, with soil of the paddock as the most likely source of infection caused by shedding of prions from the scrapie-infected sheep in this paddock up to a week earlier. 

This study also indicated that the level of contamination of field furniture sufficient to cause infection was dependent on two factors: stage of incubation period and time of last use by scrapie-infected sheep. 

Drinking from a water trough that had been used by scrapie sheep in the predominantly pre-clinical phase did not appear to cause infection, whereas infection was shown in sheep drinking from the water trough used by scrapie sheep in the later stage of the disease. 

It is possible that contamination occurred through shedding of prions in saliva, which may have contaminated the surface of the water trough and subsequently the water when it was refilled. 

Contamination appeared to be sufficient to cause infection only if the trough was in contact with sheep that included clinical cases. 

Indeed, there is an increased risk of bodily fluid infectivity with disease progression in scrapie (24) and CWD (25) based on PrPSc detection by sPMCA. 

Although ultraviolet light and heat under natural conditions do not inactivate prions (26), furniture in contact with the scrapie flock, which was assumed to be sufficiently contaminated to cause infection, did not act as vector for disease if not used for 18 months, which suggest that the weathering process alone was sufficient to inactivate prions. 

PrPSc detection by sPMCA is increasingly used as a surrogate for infectivity measurements by bioassay in sheep or mice. 

In this reported study, however, the levels of PrPSc present in the environment were below the limit of detection of the sPMCA method, yet were still sufficient to cause infection of in-contact animals. 

In the present study, the outdoor objects were removed from the infected flock 8 weeks prior to sampling and were positive by sPMCA at very low levels (2 out of 37 reactions). 

As this sPMCA assay also yielded 2 positive reactions out of 139 in samples from the scrapie-free farm, the sPMCA assay could not detect PrPSc on any of the objects above the background of the assay. 

False positive reactions with sPMCA at a low frequency associated with de novo formation of infectious prions have been reported (27, 28). 

This is in contrast to our previous study where we demonstrated that outdoor objects that had been in contact with the scrapie-infected flock up to 20 days prior to sampling harbored PrPSc that was detectable by sPMCA analysis [4 out of 15 reactions (12)] and was significantly more positive by the assay compared to analogous samples from the scrapie-free farm. 

This discrepancy could be due to the use of a different sPMCA substrate between the studies that may alter the efficiency of amplification of the environmental PrPSc. 

In addition, the present study had a longer timeframe between the objects being in contact with the infected flock and sampling, which may affect the levels of extractable PrPSc. 

Alternatively, there may be potentially patchy contamination of this furniture with PrPSc, which may have been missed by swabbing. 

The failure of sPMCA to detect CWD-associated PrP in saliva from clinically affected deer despite confirmation of infectivity in saliva-inoculated transgenic mice was associated with as yet unidentified inhibitors in saliva (29), and it is possible that the sensitivity of sPMCA is affected by other substances in the tested material. 

In addition, sampling of amplifiable PrPSc and subsequent detection by sPMCA may be more difficult from furniture exposed to weather, which is supported by the observation that PrPSc was detected by sPMCA more frequently in indoor than outdoor furniture (12). 

A recent experimental study has demonstrated that repeated cycles of drying and wetting of prion-contaminated soil, equivalent to what is expected under natural weathering conditions, could reduce PMCA amplification efficiency and extend the incubation period in hamsters inoculated with soil samples (30). 

This seems to apply also to this study even though the reduction in infectivity was more dramatic in the sPMCA assays than in the sheep model. 

Sheep were not kept until clinical end-point, which would have enabled us to compare incubation periods, but the lack of infection in sheep exposed to furniture that had not been in contact with scrapie sheep for a longer time period supports the hypothesis that prion degradation and subsequent loss of infectivity occurs even under natural conditions. 

In conclusion, the results in the current study indicate that removal of furniture that had been in contact with scrapie-infected animals should be recommended, particularly since cleaning and decontamination may not effectively remove scrapie infectivity (31), even though infectivity declines considerably if the pasture and the field furniture have not been in contact with scrapie-infected sheep for several months. As sPMCA failed to detect PrPSc in furniture that was subjected to weathering, even though exposure led to infection in sheep, this method may not always be reliable in predicting the risk of scrapie infection through environmental contamination. 

These results suggest that the VRQ/VRQ sheep model may be more sensitive than sPMCA for the detection of environmentally associated scrapie, and suggest that extremely low levels of scrapie contamination are able to cause infection in susceptible sheep genotypes. 

Keywords: classical scrapie, prion, transmissible spongiform encephalopathy, sheep, field furniture, reservoir, serial protein misfolding cyclic amplification 


**> CHRONIC WASTING DISEASE TSE PRP HUMANS ZOONOSIS ZOONOTIC <***

WHAT WE HAVE HERE, IS A LACK OF COMMUNICATION!

seems to me we might have another zoonotic tse prion disease, OR multiple new tse prion zoonotic diseases, that no one wants to talk about, and that's bad...terry

i thought i might share some news about cwd zoonosis that i got, that i cannot share or post to the public yet, i promised for various reasons, one that it will cause a shit storm for sure, but it was something i really already knew from previous studies, but, i was told that ;

==================

''As you can imagine, 2 and 5 (especially 5) may raise alarms.  The evidence we have for 4 are not as strong or tight as I would like to have.   At this point, please do not post any of the points publicly yet, but you can refer to points 1-3 in private discussions and all 5 points when discussing with relevant public officials to highlight the long-term risks of CWD zoonosis.''

====================

so, i figure your as about as official as it gets, and i think this science is extremely important for you to know and to converse about with your officials. it's about to burn a whole in my pocket. this is about as close as it will ever get for cwd zoonosis to be proven in my time, this and what Canada Czub et al found with the Macaques, plus an old study from cjd surveillance unit back that showed cjd and a 9% increase in risk from folks that eat venison, i will post all this below for your files Sir. i remember back in the BSE nvCJD days, from when the first BSE case in bovine was confirmed around 1984 maybe 83, i forget the good vets named that screwed it up first, Carol something, but from 83ish to 95 96 when nvCJD was linked to humans from BSE in cattle, so that took 10 to 15 years. hell, at that rate, especially with Texas and cwd zoonsis, hell, i'll be dead before it's official, if ever, so here ya go Sir. there was a grant study on cwd zoonosis that had been going on for some time, i followed it over the years, then the grant date for said study had expired, so, i thought i would write the good Professor about said study i.e. Professor Kong, CWRU et al. i will post the grant study abstract first, and then after that, what reply i got back, about said study that i was told not to post/publish...

CWD ZOONOSIS GRANT FIRST;

===============

Cervid to human prion transmission

Kong, Qingzhong 

Case Western Reserve University, Cleveland, OH, United States

 Abstract Prion disease is transmissible and invariably fatal. Chronic wasting disease (CWD) is the prion disease affecting deer, elk and moose, and it is a widespread and expanding epidemic affecting 22 US States and 2 Canadian provinces so far. CWD poses the most serious zoonotic prion transmission risks in North America because of huge venison consumption (>6 million deer/elk hunted and consumed annually in the USA alone), significant prion infectivity in muscles and other tissues/fluids from CWD-affected cervids, and usually high levels of individual exposure to CWD resulting from consumption of the affected animal among often just family and friends. However, we still do not know whether CWD prions can infect humans in the brain or peripheral tissues or whether clinical/asymptomatic CWD zoonosis has already occurred, and we have no essays to reliably detect CWD infection in humans. We hypothesize that: (1) The classic CWD prion strain can infect humans at low levels in the brain and peripheral lymphoid tissues; (2) The cervid-to-human transmission barrier is dependent on the cervid prion strain and influenced by the host (human) prion protein (PrP) primary sequence; (3) Reliable essays can be established to detect CWD infection in humans; and (4) CWD transmission to humans has already occurred. We will test these hypotheses in 4 Aims using transgenic (Tg) mouse models and complementary in vitro approaches. 

Aim 1 will prove that the classical CWD strain may infect humans in brain or peripheral lymphoid tissues at low levels by conducting systemic bioassays in a set of humanized Tg mouse lines expressing common human PrP variants using a number of CWD isolates at varying doses and routes. Experimental human CWD samples will also be generated for Aim 3. 

Aim 2 will test the hypothesis that the cervid-to-human prion transmission barrier is dependent on prion strain and influenced by the host (human) PrP sequence by examining and comparing the transmission efficiency and phenotypes of several atypical/unusual CWD isolates/strains as well as a few prion strains from other species that have adapted to cervid PrP sequence, utilizing the same panel of humanized Tg mouse lines as in Aim 1. 

Aim 3 will establish reliable essays for detection and surveillance of CWD infection in humans by examining in details the clinical, pathological, biochemical and in vitro seeding properties of existing and future experimental human CWD samples generated from Aims 1-2 and compare them with those of common sporadic human Creutzfeldt-Jakob disease (sCJD) prions. 

Aim 4 will attempt to detect clinical CWD-affected human cases by examining a significant number of brain samples from prion-affected human subjects in the USA and Canada who have consumed venison from CWD-endemic areas utilizing the criteria and essays established in Aim 3. The findings from this proposal will greatly advance our understandings on the potential and characteristics of cervid prion transmission in humans, establish reliable essays for CWD zoonosis and potentially discover the first case(s) of CWD infection in humans.

Public Health Relevance There are significant and increasing human exposure to cervid prions because chronic wasting disease (CWD, a widespread and highly infectious prion disease among deer and elk in North America) continues spreading and consumption of venison remains popular, but our understanding on cervid-to-human prion transmission is still very limited, raising public health concerns. This proposal aims to define the zoonotic risks of cervid prions and set up and apply essays to detect CWD zoonosis using mouse models and in vitro methods. The findings will greatly expand our knowledge on the potentials and characteristics of cervid prion transmission in humans, establish reliable essays for such infections and may discover the first case(s) of CWD infection in humans.

 Funding Agency Agency National Institute of Health (NIH) Institute National Institute of Neurological Disorders and Stroke (NINDS) Type Research Project (R01) Project # 1R01NS088604-01A1 Application # 9037884 Study Section Cellular and Molecular Biology of Neurodegeneration Study Section (CMND) Program Officer Wong, May Project Start 2015-09-30 Project End 2019-07-31 Budget Start 2015-09-30 Budget End 2016-07-31 Support Year 1 Fiscal Year 2015 Total Cost $337,507 Indirect Cost $118,756

snip... 


Professor Kongs reply to me just this month about above grant study that has NOT been published in peer reveiw yet...

=================================

Here is a brief summary of our findings:

snip...can't post, made a promise...tss

On Sat, Apr 3, 2021 at 12:19 PM Terry Singeltary <flounder9@verizon.net> wrote:

snip...

end...tss

==============

CWD ZOONOSIS THE FULL MONTY TO DATE

International Conference on Emerging Diseases, Outbreaks & Case Studies & 16th Annual Meeting on Influenza March 28-29, 2018 | Orlando, USA

Qingzhong Kong

Case Western Reserve University School of Medicine, USA

Zoonotic potential of chronic wasting disease prions from cervids

Chronic wasting disease (CWD) is the prion disease in cervids (mule deer, white-tailed deer, American elk, moose, and reindeer). It has become an epidemic in North America, and it has been detected in the Europe (Norway) since 2016. The widespread CWD and popular hunting and consumption of cervid meat and other products raise serious public health concerns, but questions remain on human susceptibility to CWD prions, especially on the potential difference in zoonotic potential among the various CWD prion strains. We have been working to address this critical question for well over a decade. We used CWD samples from various cervid species to inoculate transgenic mice expressing human or elk prion protein (PrP). We found infectious prions in the spleen or brain in a small fraction of CWD-inoculated transgenic mice expressing human PrP, indicating that humans are not completely resistant to CWD prions; this finding has significant ramifications on the public health impact of CWD prions. The influence of cervid PrP polymorphisms, the prion strain dependence of CWD-to-human transmission barrier, and the characterization of experimental human CWD prions will be discussed.

Speaker Biography Qingzhong Kong has completed his PhD from the University of Massachusetts at Amherst and Post-doctoral studies at Yale University. He is currently an Associate Professor of Pathology, Neurology and Regenerative Medicine. He has published over 50 original research papers in reputable journals (including Science Translational Medicine, JCI, PNAS and Cell Reports) and has been serving as an Editorial Board Member on seven scientific journals. He has multiple research interests, including public health risks of animal prions (CWD of cervids and atypical BSE of cattle), animal modeling of human prion diseases, mechanisms of prion replication and pathogenesis, etiology of sporadic Creutzfeldt-Jacob disease (CJD) in humans, normal cellular PrP in the biology and pathology of multiple brain and peripheral diseases, proteins responsible for the α-cleavage of cellular PrP, as well as gene therapy and DNA vaccination.






SUNDAY, JULY 25, 2021 

North American and Norwegian Chronic Wasting Disease prions exhibit different potential for interspecies transmission and zoonotic risk 

''Our data suggest that reindeer and red deer from Norway could be the most transmissible CWD prions to other mammals, whereas North American CWD prions were more prone to generate human prions in vitro.''


MONDAY, JULY 19, 2021 

***> U Calgary researchers at work on a vaccine against a fatal infectious disease affecting deer and potentially people


Prion Conference 2018 Abstracts

BSE aka MAD COW DISEASE, was first discovered in 1984, and it took until 1995 to finally admit that BSE was causing nvCJD, the rest there is history, but that science is still evolving i.e. science now shows that indeed atypical L-type BSE, atypical Nor-98 Scrapie, and typical Scrapie are all zoonosis, zoonotic for humans, there from. 

HOW long are we going to wait for Chronic Wasting Disease, CWD TSE Prion of Cervid, and zoonosis, zoonotic tranmission to humans there from?

Studies have shown since 1994 that humans are susceptible to CWD TSE Prion, so, what's the hold up with making CWD a zoonotic zoonosis disease, the iatrogenic transmissions there from is not waiting for someone to make a decision.

Prion Conference 2018 Abstracts

P190 Human prion disease mortality rates by occurrence of chronic wasting disease in freeranging cervids, United States

Abrams JY (1), Maddox RA (1), Schonberger LB (1), Person MK (1), Appleby BS (2), Belay ED (1)

(1) Centers for Disease Control and Prevention (CDC), National Center for Emerging and Zoonotic Infectious Diseases, Atlanta, GA, USA (2) Case Western Reserve University, National Prion Disease Pathology Surveillance Center (NPDPSC), Cleveland, OH, USA.

Background

Chronic wasting disease (CWD) is a prion disease of deer and elk that has been identified in freeranging cervids in 23 US states. While there is currently no epidemiological evidence for zoonotic transmission through the consumption of contaminated venison, studies suggest the CWD agent can cross the species barrier in experimental models designed to closely mimic humans. We compared rates of human prion disease in states with and without CWD to examine the possibility of undetermined zoonotic transmission.

Methods

Death records from the National Center for Health Statistics, case records from the National Prion Disease Pathology Surveillance Center, and additional state case reports were combined to create a database of human prion disease cases from 2003-2015. Identification of CWD in each state was determined through reports of positive CWD tests by state wildlife agencies. Age- and race-adjusted mortality rates for human prion disease, excluding cases with known etiology, were determined for four categories of states based on CWD occurrence: highly endemic (>16 counties with CWD identified in free-ranging cervids); moderately endemic (3-10 counties with CWD); low endemic (1-2 counties with CWD); and no CWD states. States were counted as having no CWD until the year CWD was first identified. Analyses stratified by age, sex, and time period were also conducted to focus on subgroups for which zoonotic transmission would be more likely to be detected: cases <55 years old, male sex, and the latter half of the study (2010-2015).

Results

Highly endemic states had a higher rate of prion disease mortality compared to non-CWD states (rate ratio [RR]: 1.12, 95% confidence interval [CI] = 1.01 - 1.23), as did low endemic states (RR: 1.15, 95% CI = 1.04 - 1.27). Moderately endemic states did not have an elevated mortality rate (RR: 1.05, 95% CI = 0.93 - 1.17). In age-stratified analyses, prion disease mortality rates among the <55 year old population were elevated for moderately endemic states (RR: 1.57, 95% CI = 1.10 – 2.24) while mortality rates were elevated among those ≥55 for highly endemic states (RR: 1.13, 95% CI = 1.02 - 1.26) and low endemic states (RR: 1.16, 95% CI = 1.04 - 1.29). In other stratified analyses, prion disease mortality rates for males were only elevated for low endemic states (RR: 1.27, 95% CI = 1.10 - 1.48), and none of the categories of CWD-endemic states had elevated mortality rates for the latter time period (2010-2015).

Conclusions

While higher prion disease mortality rates in certain categories of states with CWD in free-ranging cervids were noted, additional stratified analyses did not reveal markedly elevated rates for potentially sensitive subgroups that would be suggestive of zoonotic transmission. Unknown confounding factors or other biases may explain state-by-state differences in prion disease mortality.

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P172 Peripheral Neuropathy in Patients with Prion Disease

Wang H(1), Cohen M(1), Appleby BS(1,2)

(1) University Hospitals Cleveland Medical Center, Cleveland, Ohio (2) National Prion Disease Pathology Surveillance Center, Cleveland, Ohio.

Prion disease is a fatal progressive neurodegenerative disease due to deposition of an abnormal protease-resistant isoform of prion protein. Typical symptoms include rapidly progressive dementia, myoclonus, visual disturbance and hallucinations. Interestingly, in patients with prion disease, the abnormal protein canould also be found in the peripheral nervous system. Case reports of prion deposition in peripheral nerves have been reported. Peripheral nerve involvement is thought to be uncommon; however, little is known about the exact prevalence and features of peripheral neuropathy in patients with prion disease.

We reviewed autopsy-proven prion cases from the National Prion Disease Pathology Surveillance Center that were diagnosed between September 2016 to March 2017. We collected information regarding prion protein diagnosis, demographics, comorbidities, clinical symptoms, physical exam, neuropathology, molecular subtype, genetics lab, brain MRI, image and EMG reports. Our study included 104 patients. Thirteen (12.5%) patients had either subjective symptoms or objective signs of peripheral neuropathy. Among these 13 patients, 3 had other known potential etiologies of peripheral neuropathy such as vitamin B12 deficiency or prior chemotherapy. Among 10 patients that had no other clear etiology, 3 (30%) had familial CJD. The most common sCJD subtype was MV1-2 (30%), followed by MM1-2 (20%). The Majority of cases wasere male (60%). Half of them had exposure to wild game. The most common subjective symptoms were tingling and/or numbness of distal extremities. The most common objective finding was diminished vibratory sensation in the feet. Half of them had an EMG with the findings ranging from fasciculations to axonal polyneuropathy or demyelinating polyneuropathy.

Our study provides an overview of the pattern of peripheral neuropathy in patients with prion disease. Among patients with peripheral neuropathy symptoms or signs, majority has polyneuropathy. It is important to document the baseline frequency of peripheral neuropathy in prion diseases as these symptoms may become important when conducting surveillance for potential novel zoonotic prion diseases.

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P177 PrP plaques in methionine homozygous Creutzfeldt-Jakob disease patients as a potential marker of iatrogenic transmission

Abrams JY (1), Schonberger LB (1), Cali I (2), Cohen Y (2), Blevins JE (2), Maddox RA (1), Belay ED (1), Appleby BS (2), Cohen ML (2)

(1) Centers for Disease Control and Prevention (CDC), National Center for Emerging and Zoonotic Infectious Diseases, Atlanta, GA, USA (2) Case Western Reserve University, National Prion Disease Pathology Surveillance Center (NPDPSC), Cleveland, OH, USA.

Background

Sporadic Creutzfeldt-Jakob disease (CJD) is widely believed to originate from de novo spontaneous conversion of normal prion protein (PrP) to its pathogenic form, but concern remains that some reported sporadic CJD cases may actually be caused by disease transmission via iatrogenic processes. For cases with methionine homozygosity (CJD-MM) at codon 129 of the PRNP gene, recent research has pointed to plaque-like PrP deposition as a potential marker of iatrogenic transmission for a subset of cases. This phenotype is theorized to originate from specific iatrogenic source CJD types that comprise roughly a quarter of known CJD cases.

Methods

We reviewed scientific literature for studies which described PrP plaques among CJD patients with known epidemiological links to iatrogenic transmission (receipt of cadaveric human grown hormone or dura mater), as well as in cases of reported sporadic CJD. The presence and description of plaques, along with CJD classification type and other contextual factors, were used to summarize the current evidence regarding plaques as a potential marker of iatrogenic transmission. In addition, 523 cases of reported sporadic CJD cases in the US from January 2013 through September 2017 were assessed for presence of PrP plaques.

Results

We identified four studies describing 52 total cases of CJD-MM among either dura mater recipients or growth hormone recipients, of which 30 were identified as having PrP plaques. While sporadic cases were not generally described as having plaques, we did identify case reports which described plaques among sporadic MM2 cases as well as case reports of plaques exclusively in white matter among sporadic MM1 cases. Among the 523 reported sporadic CJD cases, 0 of 366 MM1 cases had plaques, 2 of 48 MM2 cases had kuru plaques, and 4 of 109 MM1+2 cases had either kuru plaques or both kuru and florid plaques. Medical chart review of the six reported sporadic CJD cases with plaques did not reveal clinical histories suggestive of potential iatrogenic transmission.

Conclusions

PrP plaques occur much more frequently for iatrogenic CJD-MM cases compared to sporadic CJDMM cases. Plaques may indicate iatrogenic transmission for CJD-MM cases without a type 2 Western blot fragment. The study results suggest the absence of significant misclassifications of iatrogenic CJD as sporadic. To our knowledge, this study is the first to describe grey matter kuru plaques in apparently sporadic CJD-MM patients with a type 2 Western blot fragment.

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P180 Clinico-pathological analysis of human prion diseases in a brain bank series

Ximelis T (1), Aldecoa I (1,2), Molina-Porcel L (1,3), Grau-Rivera O (4), Ferrer I (5), Nos C (6), Gelpi E (1,7), Sánchez-Valle R (1,4)

(1) Neurological Tissue Bank of the Biobanc-Hospital ClÃnic-IDIBAPS, Barcelona, Spain (2) Pathological Service of Hospital ClÃnic de Barcelona, Barcelona, Spain (3) EAIA Trastorns Cognitius, Centre Emili Mira, Parc de Salut Mar, Barcelona, Spain (4) Department of Neurology of Hospital ClÃnic de Barcelona, Barcelona, Spain (5) Institute of Neuropathology, Hospital Universitari de Bellvitge, Hospitalet de Llobregat, Barcelona (6) General subdirectorate of Surveillance and Response to Emergencies in Public Health, Department of Public Health in Catalonia, Barcelona, Spain (7) Institute of Neurology, Medical University of Vienna, Vienna, Austria.

Background and objective:

The Neurological Tissue Bank (NTB) of the Hospital Clínic-Institut d‘Investigacions Biomèdiques August Pi i Sunyer, Barcelona, Spain is the reference center in Catalonia for the neuropathological study of prion diseases in the region since 2001. The aim of this study is to analyse the characteristics of the confirmed prion diseases registered at the NTB during the last 15 years.

Methods:

We reviewed retrospectively all neuropathologically confirmed cases registered during the period January 2001 to December 2016.

Results:

176 cases (54,3% female, mean age: 67,5 years and age range: 25-86 years) of neuropathological confirmed prion diseases have been studied at the NTB. 152 cases corresponded to sporadic Creutzfeldt-Jakob disease (sCJD), 10 to genetic CJD, 10 to Fatal Familial Insomnia, 2 to GerstmannSträussler-Scheinker disease, and 2 cases to variably protease-sensitive prionopathy (VPSPr). Within sCJD subtypes the MM1 subtype was the most frequent, followed by the VV2 histotype.

Clinical and neuropathological diagnoses agreed in 166 cases (94%). The clinical diagnosis was not accurate in 10 patients with definite prion disease: 1 had a clinical diagnosis of Fronto-temporal dementia (FTD), 1 Niemann-Pick‘s disease, 1 Lewy Body‘s Disease, 2 Alzheimer‘s disease, 1 Cortico-basal syndrome and 2 undetermined dementia. Among patients with VPSPr, 1 had a clinical diagnosis of Amyotrophic lateral sclerosis (ALS) and the other one with FTD.

Concomitant pathologies are frequent in older age groups, mainly AD neuropathological changes were observed in these subjects.

Discussion:

A wide spectrum of human prion diseases have been identified in the NTB being the relative frequencies and main characteristics like other published series. There is a high rate of agreement between clinical and neuropathological diagnoses with prion diseases. These findings show the importance that public health has given to prion diseases during the past 15 years. Continuous surveillance of human prion disease allows identification of new emerging phenotypes. Brain tissue samples from these donors are available to the scientific community. For more information please visit:


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P192 Prion amplification techniques for the rapid evaluation of surface decontamination procedures

Bruyere-Ostells L (1), Mayran C (1), Belondrade M (1), Boublik Y (2), Haïk S (3), Fournier-Wirth C (1), Nicot S (1), Bougard D (1)

(1) Pathogenesis and control of chronic infections, Etablissement Français du Sang, Inserm, Université de Montpellier, Montpellier, France. (2) Centre de Recherche en Biologie cellulaire de Montpellier, CNRS, Université de Montpellier, Montpellier, France. (3) Inserm U 1127, CNRS UMR 7225, Sorbonne Universités, UPMC Université Paris 06 UMR S 1127, Institut du Cerveau et de la Moelle épinière, ICM, Paris, France.

Aims:

Transmissible Spongiform Encephalopathies (TSE) or prion diseases are a group of incurable and always fatal neurodegenerative disorders including Creutzfeldt-Jakob diseases (CJD) in humans. These pathologies include sporadic (sCJD), genetic and acquired (variant CJD) forms. By the past, sCJD and vCJD were transmitted by different prion contaminated biological materials to patients resulting in more than 400 iatrogenic cases (iCJD). The atypical nature and the biochemical properties of the infectious agent, formed by abnormal prion protein or PrPTSE, make it particularly resistant to conventional decontamination procedures. In addition, PrPTSE is widely distributed throughout the organism before clinical onset in vCJD and can also be detected in some peripheral tissues in sporadic CJD. Risk of iatrogenic transmission of CJD by contaminated medical device remains thus a concern for healthcare facilities. Bioassay is the gold standard method to evaluate the efficacy of prion decontamination procedures but is time-consuming and expensive. Here, we propose to compare in vitro prion amplification techniques: Protein Misfolding Cyclic Amplification (PMCA) and Real-Time Quaking Induced Conversion (RT-QuIC) for the detection of residual prions on surface after decontamination.

Methods:

Stainless steel wires, by mimicking the surface of surgical instruments, were proposed as a carrier model of prions for inactivation studies. To determine the sensitivity of the two amplification techniques on wires (Surf-PMCA and Surf-QuIC), steel wires were therefore contaminated with serial dilutions of brain homogenates (BH) from a 263k infected hamster and from a patient with sCJD (MM1 subtype). We then compared the different standard decontamination procedures including partially and fully efficient treatments by detecting the residual seeding activity on 263K and sCJD contaminated wires. We completed our study by the evaluation of marketed reagents endorsed for prion decontamination.

Results:

The two amplification techniques can detect minute quantities of PrPTSE adsorbed onto a single wire. 8/8 wires contaminated with a 10-6 dilution of 263k BH and 1/6 with the 10-8 dilution are positive with Surf-PMCA. Similar performances were obtained with Surf-QuIC on 263K: 10/16 wires contaminated with 10-6 dilution and 1/8 wires contaminated with 10-8 dilution are positive. Regarding the human sCJD-MM1 prion, Surf-QuIC allows us to detect 16/16 wires contaminated with 10-6 dilutions and 14/16 with 10-7 . Results obtained after decontamination treatments are very similar between 263K and sCJD prions. Efficiency of marketed treatments to remove prions is lower than expected.

Conclusions:

Surf-PMCA and Surf-QuIC are very sensitive methods for the detection of prions on wires and could be applied to prion decontamination studies for rapid evaluation of new treatments. Sodium hypochlorite is the only product to efficiently remove seeding activity of both 263K and sCJD prions.

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WA2 Oral transmission of CWD into Cynomolgus macaques: signs of atypical disease, prion conversion and infectivity in macaques and bio-assayed transgenic mice

Schatzl HM (1, 2), Hannaoui S (1, 2), Cheng Y-C (1, 2), Gilch S (1, 2), Beekes M (3), SchulzSchaeffer W (4), Stahl-Hennig C (5) and Czub S (2, 6)

(1) University of Calgary, Calgary Prion Research Unit, Calgary, Canada (2) University of Calgary, Faculty of Veterinary Medicine, Calgary, Canada, (3) Robert Koch Institute, Berlin, Germany, (4) University of Homburg/Saar, Homburg, Germany, (5) German Primate Center, Goettingen, Germany, (6) Canadian Food Inspection Agency (CFIA), Lethbridge, Canada.

To date, BSE is the only example of interspecies transmission of an animal prion disease into humans. The potential zoonotic transmission of CWD is an alarming issue and was addressed by many groups using a variety of in vitro and in vivo experimental systems. Evidence from these studies indicated a substantial, if not absolute, species barrier, aligning with the absence of epidemiological evidence suggesting transmission into humans. Studies in non-human primates were not conclusive so far, with oral transmission into new-world monkeys and no transmission into old-world monkeys. Our consortium has challenged 18 Cynomolgus macaques with characterized CWD material, focusing on oral transmission with muscle tissue. Some macaques have orally received a total of 5 kg of muscle material over a period of 2 years. After 5-7 years of incubation time some animals showed clinical symptoms indicative of prion disease, and prion neuropathology and PrPSc deposition were found in spinal cord and brain of euthanized animals. PrPSc in immunoblot was weakly detected in some spinal cord materials and various tissues tested positive in RT-QuIC, including lymph node and spleen homogenates. To prove prion infectivity in the macaque tissues, we have intracerebrally inoculated 2 lines of transgenic mice, expressing either elk or human PrP. At least 3 TgElk mice, receiving tissues from 2 different macaques, showed clinical signs of a progressive prion disease and brains were positive in immunoblot and RT-QuIC. Tissues (brain, spinal cord and spleen) from these and preclinical mice are currently tested using various read-outs and by second passage in mice. Transgenic mice expressing human PrP were so far negative for clear clinical prion disease (some mice >300 days p.i.). In parallel, the same macaque materials are inoculated into bank voles. Taken together, there is strong evidence of transmissibility of CWD orally into macaques and from macaque tissues into transgenic mouse models, although with an incomplete attack rate. The clinical and pathological presentation in macaques was mostly atypical, with a strong emphasis on spinal cord pathology. Our ongoing studies will show whether the transmission of CWD into macaques and passage in transgenic mice represents a form of non-adaptive prion amplification, and whether macaque-adapted prions have the potential to infect mice expressing human PrP. The notion that CWD can be transmitted orally into both new-world and old-world non-human primates asks for a careful reevaluation of the zoonotic risk of CWD.

See also poster P103

***> The notion that CWD can be transmitted orally into both new-world and old-world non-human primates asks for a careful reevaluation of the zoonotic risk of CWD.

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WA16 Monitoring Potential CWD Transmission to Humans

Belay ED

Centers for Disease Control and Prevention (CDC), National Center for Emerging and Zoonotic Infectious Diseases, Atlanta, GA, USA.

The spread of chronic wasting disease (CWD) in animals has raised concerns about increasing human exposure to the CWD agent via hunting and venison consumption, potentially facilitating CWD transmission to humans. Several studies have explored this possibility, including limited epidemiologic studies, in vitro experiments, and laboratory studies using various types of animal models. Most human exposures to the CWD agent in the United States would be expected to occur in association with deer and elk hunting in CWD-endemic areas. The Centers for Disease Control and Prevention (CDC) collaborated with state health departments in Colorado, Wisconsin, and Wyoming to identify persons at risk of CWD exposure and to monitor their vital status over time. Databases were established of persons who hunted in Colorado and Wyoming and those who reported consumption of venison from deer that later tested positive in Wisconsin. Information from the databases is periodically cross-checked with mortality data to determine the vital status and causes of death for deceased persons. Long-term follow-up of these hunters is needed to assess their risk of development of a prion disease linked to CWD exposure.

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P166 Characterization of CJD strain profiles in venison consumers and non-consumers from Alberta and Saskatchewan

Stephanie Booth (1,2), Lise Lamoureux (1), Debra Sorensen (1), Jennifer L. Myskiw (1,2), Megan Klassen (1,2), Michael Coulthart (3), Valerie Sim (4)

(1) Zoonotic Diseases and Special Pathogens, National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg (2) Department of Medical Microbiology and Infectious Diseases, University of Manitoba, Winnipeg (3) Canadian CJD Surveillance System, Public Health Agency of Canada, Ottawa (4) Division of Neurology, Department of Medicine Centre for Prions and Protein Folding Diseases, University of Alberta, Edmonton.

Chronic wasting disease (CWD) is spreading rapidly through wild cervid populations in the Canadian provinces of Alberta and Saskatchewan. While this has implications for tourism and hunting, there is also concern over possible zoonotic transmission to humans who eat venison from infected deer. Whilst there is no evidence of any human cases of CWD to date, the Canadian CJD Surveillance System (CJDSS) in Canada is staying vigilant. When variant CJD occurred following exposure to BSE, the unique biochemical fingerprint of the pathologic PrP enabled a causal link to be confirmed. However, we cannot be sure what phenotype human CWD prions would present with, or indeed, whether this would be distinct from that see in sporadic CJD. Therefore we are undertaking a systematic analysis of the molecular diversity of CJD cases of individuals who resided in Alberta and Saskatchewan at their time of death comparing venison consumers and non-consumers, using a variety of clinical, imaging, pathological and biochemical markers. Our initial objective is to develop novel biochemical methodologies that will extend the baseline glycoform and genetic polymorphism typing that is already completed by the CJDSS. Firstly, we are reviewing MRI, EEG and pathology information from over 40 cases of CJD to select clinically affected areas for further investigation. Biochemical analysis will include assessment of the levels of protease sensitive and resistant prion protein, glycoform typing using 2D gel electrophoresis, testing seeding capabilities and kinetics of aggregation by quaking-induced conversion, and determining prion oligomer size distributions with asymmetric flow field fractionation with in-line light scattering. Progress and preliminary data will be presented. Ultimately, we intend to further define the relationship between PrP structure and disease phenotype and establish a baseline for the identification of future atypical CJD cases that may arise as a result of exposure to CWD.

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Source Prion Conference 2018 Abstracts




Volume 24, Number 8—August 2018 Research Susceptibility of Human Prion Protein to Conversion by Chronic Wasting Disease Prions

Marcelo A. BarriaComments to Author , Adriana Libori, Gordon Mitchell, and Mark W. Head Author affiliations: National CJD Research and Surveillance Unit, University of Edinburgh, Edinburgh, Scotland, UK (M.A. Barria, A. Libori, M.W. Head); National and OIE Reference Laboratory for Scrapie and CWD, Canadian Food Inspection Agency, Ottawa, Ontario, Canada (G. Mitchell)

Abstract Chronic wasting disease (CWD) is a contagious and fatal neurodegenerative disease and a serious animal health issue for deer and elk in North America. The identification of the first cases of CWD among free-ranging reindeer and moose in Europe brings back into focus the unresolved issue of whether CWD can be zoonotic like bovine spongiform encephalopathy. We used a cell-free seeded protein misfolding assay to determine whether CWD prions from elk, white-tailed deer, and reindeer in North America can convert the human prion protein to the disease-associated form. We found that prions can convert, but the efficiency of conversion is affected by polymorphic variation in the cervid and human prion protein genes. In view of the similarity of reindeer, elk, and white-tailed deer in North America to reindeer, red deer, and roe deer, respectively, in Europe, a more comprehensive and thorough assessment of the zoonotic potential of CWD might be warranted.

snip...

Discussion Characterization of the transmission properties of CWD and evaluation of their zoonotic potential are important for public health purposes. Given that CWD affects several members of the family Cervidae, it seems reasonable to consider whether the zoonotic potential of CWD prions could be affected by factors such as CWD strain, cervid species, geographic location, and Prnp–PRNP polymorphic variation. We have previously used an in vitro conversion assay (PMCA) to investigate the susceptibility of the human PrP to conversion to its disease-associated form by several animal prion diseases, including CWD (15,16,22). The sensitivity of our molecular model for the detection of zoonotic conversion depends on the combination of 1) the action of proteinase K to degrade the abundant human PrPC that constitutes the substrate while only N terminally truncating any human PrPres produced and 2) the presence of the 3F4 epitope on human but not cervid PrP. In effect, this degree of sensitivity means that any human PrPres formed during the PMCA reaction can be detected down to the limit of Western blot sensitivity. In contrast, if other antibodies that detect both cervid and human PrP are used, such as 6H4, then newly formed human PrPres must be detected as a measurable increase in PrPres over the amount remaining in the reaction product from the cervid seed. Although best known for the efficient amplification of prions in research and diagnostic contexts, the variation of the PMCA method employed in our study is optimized for the definitive detection of zoonotic reaction products of inherently inefficient conversion reactions conducted across species barriers. By using this system, we previously made and reported the novel observation that elk CWD prions could convert human PrPC from human brain and could also convert recombinant human PrPC expressed in transgenic mice and eukaryotic cell cultures (15).

A previous publication suggested that mule deer PrPSc was unable to convert humanized transgenic substrate in PMCA assays (23) and required a further step of in vitro conditioning in deer substrate PMCA before it was able to cross the deer–human molecular barrier (24). However, prions from other species, such as elk (15) and reindeer affected by CWD, appear to be compatible with the human protein in a single round of amplification (as shown in our study). These observations suggest that different deer species affected by CWD could present differing degrees of the olecular compatibility with the normal form of human PrP.

The contribution of the polymorphism at codon 129 of the human PrP gene has been extensively studied and is recognized as a risk factor for Creutzfeldt-Jakob disease (4). In cervids, the equivalent codon corresponds to the position 132 encoding methionine or leucine. This polymorphism in the elk gene has been shown to play an important role in CWD susceptibility (25,26). We have investigated the effect of this cervid Prnp polymorphism on the conversion of the humanized transgenic substrate according to the variation in the equivalent PRNP codon 129 polymorphism. Interestingly, only the homologs methionine homozygous seed–substrate reactions could readily convert the human PrP, whereas the heterozygous elk PrPSc was unable to do so, even though comparable amounts of PrPres were used to seed the reaction. In addition, we observed only low levels of human PrPres formation in the reactions seeded with the homozygous methionine (132 MM) and the heterozygous (132 ML) seeds incubated with the other 2 human polymorphic substrates (129 MV and 129 VV). The presence of the amino acid leucine at position 132 of the elk Prnp gene has been attributed to a lower degree of prion conversion compared with methionine on the basis of experiments in mice made transgenic for these polymorphic variants (26). Considering the differences observed for the amplification of the homozygous human methionine substrate by the 2 polymorphic elk seeds (MM and ML), reappraisal of the susceptibility of human PrPC by the full range of cervid polymorphic variants affected by CWD would be warranted.

In light of the recent identification of the first cases of CWD in Europe in a free-ranging reindeer (R. tarandus) in Norway (2), we also decided to evaluate the in vitro conversion potential of CWD in 2 experimentally infected reindeer (18). Formation of human PrPres was readily detectable after a single round of PMCA, and in all 3 humanized polymorphic substrates (MM, MV, and VV). This finding suggests that CWD prions from reindeer could be more compatible with human PrPC generally and might therefore present a greater risk for zoonosis than, for example, CWD prions from white-tailed deer. A more comprehensive comparison of CWD in the affected species, coupled with the polymorphic variations in the human and deer PRNP–Prnp genes, in vivo and in vitro, will be required before firm conclusions can be drawn. Analysis of the Prnp sequence of the CWD reindeer in Norway was reported to be identical to the specimens used in our study (2). This finding raises the possibility of a direct comparison of zoonotic potential between CWD acquired in the wild and that produced in a controlled laboratory setting. (Table).

The prion hypothesis proposes that direct molecular interaction between PrPSc and PrPC is necessary for conversion and prion replication. Accordingly, polymorphic variants of the PrP of host and agent might play a role in determining compatibility and potential zoonotic risk. In this study, we have examined the capacity of the human PrPC to support in vitro conversion by elk, white-tailed deer, and reindeer CWD PrPSc. Our data confirm that elk CWD prions can convert the human PrPC, at least in vitro, and show that the homologous PRNP polymorphisms at codon 129 and 132 in humans and cervids affect conversion efficiency. Other species affected by CWD, particularly caribou or reindeer, also seem able to convert the human PrP. It will be important to determine whether other polymorphic variants found in other CWD-affected Cervidae or perhaps other factors (17) exert similar effects on the ability to convert human PrP and thus affect their zoonotic potential.

Dr. Barria is a research scientist working at the National CJD Research and Surveillance Unit, University of Edinburgh. His research has focused on understanding the molecular basis of a group of fatal neurologic disorders called prion diseases.

Acknowledgments We thank Aru Balachandran for originally providing cervid brain tissues, Abigail Diack and Jean Manson for providing mouse brain tissue, and James Ironside for his critical reading of the manuscript at an early stage.

This report is independent research commissioned and funded by the United Kingdom’s Department of Health Policy Research Programme and the Government of Scotland. The views expressed in this publication are those of the authors and not necessarily those of the Department of Health or the Government of Scotland.

Author contributions: The study was conceived and designed by M.A.B. and M.W.H. The experiments were conducted by M.A.B. and A.L. Chronic wasting disease brain specimens were provided by G.M. The manuscript was written by M.A.B. and M.W.H. All authors contributed to the editing and revision of the manuscript.



Prion 2017 Conference Abstracts
First evidence of intracranial and peroral transmission of Chronic Wasting Disease (CWD) into Cynomolgus macaques: a work in progress Stefanie Czub1, Walter Schulz-Schaeffer2, Christiane Stahl-Hennig3, Michael Beekes4, Hermann Schaetzl5 and Dirk Motzkus6 1 
University of Calgary Faculty of Veterinary Medicine/Canadian Food Inspection Agency; 2Universitatsklinikum des Saarlandes und Medizinische Fakultat der Universitat des Saarlandes; 3 Deutsches Primaten Zentrum/Goettingen; 4 Robert-Koch-Institut Berlin; 5 University of Calgary Faculty of Veterinary Medicine; 6 presently: Boehringer Ingelheim Veterinary Research Center; previously: Deutsches Primaten Zentrum/Goettingen 
This is a progress report of a project which started in 2009. 
21 cynomolgus macaques were challenged with characterized CWD material from white-tailed deer (WTD) or elk by intracerebral (ic), oral, and skin exposure routes. Additional blood transfusion experiments are supposed to assess the CWD contamination risk of human blood product. Challenge materials originated from symptomatic cervids for ic, skin scarification and partially per oral routes (WTD brain). Challenge material for feeding of muscle derived from preclinical WTD and from preclinical macaques for blood transfusion experiments. We have confirmed that the CWD challenge material contained at least two different CWD agents (brain material) as well as CWD prions in muscle-associated nerves. 
Here we present first data on a group of animals either challenged ic with steel wires or per orally and sacrificed with incubation times ranging from 4.5 to 6.9 years at postmortem. Three animals displayed signs of mild clinical disease, including anxiety, apathy, ataxia and/or tremor. In four animals wasting was observed, two of those had confirmed diabetes. All animals have variable signs of prion neuropathology in spinal cords and brains and by supersensitive IHC, reaction was detected in spinal cord segments of all animals. Protein misfolding cyclic amplification (PMCA), real-time quaking-induced conversion (RT-QuiC) and PET-blot assays to further substantiate these findings are on the way, as well as bioassays in bank voles and transgenic mice. 
At present, a total of 10 animals are sacrificed and read-outs are ongoing. Preclinical incubation of the remaining macaques covers a range from 6.4 to 7.10 years. Based on the species barrier and an incubation time of > 5 years for BSE in macaques and about 10 years for scrapie in macaques, we expected an onset of clinical disease beyond 6 years post inoculation. 
PRION 2017 DECIPHERING NEURODEGENERATIVE DISORDERS ABSTRACTS REFERENCE
8. Even though human TSE‐exposure risk through consumption of game from European cervids can be assumed to be minor, if at all existing, no final conclusion can be drawn due to the overall lack of scientific data. In particular the US data do not clearly exclude the possibility of human (sporadic or familial) TSE development due to consumption of venison. The Working Group thus recognizes a potential risk to consumers if a TSE would be present in European cervids. It might be prudent considering appropriate measures to reduce such a risk, e.g. excluding tissues such as CNS and lymphoid tissues from the human food chain, which would greatly reduce any potential risk for consumers. However, it is stressed that currently, no data regarding a risk of TSE infections from cervid products are available.


SATURDAY, FEBRUARY 23, 2019 

Chronic Wasting Disease CWD TSE Prion and THE FEAST 2003 CDC an updated review of the science 2019


TUESDAY, NOVEMBER 04, 2014 

Six-year follow-up of a point-source exposure to CWD contaminated venison in an Upstate New York community: risk behaviours and health outcomes 2005–2011

Authors, though, acknowledged the study was limited in geography and sample size and so it couldn't draw a conclusion about the risk to humans. They recommended more study. Dr. Ermias Belay was the report's principal author but he said New York and Oneida County officials are following the proper course by not launching a study. "There's really nothing to monitor presently. No one's sick," Belay said, noting the disease's incubation period in deer and elk is measured in years. "


Transmission Studies

Mule deer transmissions of CWD were by intracerebral inoculation and compared with natural cases {the following was written but with a single line marked through it ''first passage (by this route)}....TSS

resulted in a more rapidly progressive clinical disease with repeated episodes of synocopy ending in coma. One control animal became affected, it is believed through contamination of inoculum (?saline). Further CWD transmissions were carried out by Dick Marsh into ferret, mink and squirrel monkey. Transmission occurred in ALL of these species with the shortest incubation period in the ferret.

snip.... 


Prion Infectivity in Fat of Deer with Chronic Wasting Disease▿ 

Brent Race#, Kimberly Meade-White#, Richard Race and Bruce Chesebro* + Author Affiliations

In mice, prion infectivity was recently detected in fat. Since ruminant fat is consumed by humans and fed to animals, we determined infectivity titers in fat from two CWD-infected deer. Deer fat devoid of muscle contained low levels of CWD infectivity and might be a risk factor for prion infection of other species. 


Prions in Skeletal Muscles of Deer with Chronic Wasting Disease 

Here bioassays in transgenic mice expressing cervid prion protein revealed the presence of infectious prions in skeletal muscles of CWD-infected deer, demonstrating that humans consuming or handling meat from CWD-infected deer are at risk to prion exposure. 


*** now, let’s see what the authors said about this casual link, personal communications years ago, and then the latest on the zoonotic potential from CWD to humans from the TOKYO PRION 2016 CONFERENCE.

see where it is stated NO STRONG evidence. so, does this mean there IS casual evidence ???? “Our conclusion stating that we found no strong evidence of CWD transmission to humans”

From: TSS 

Subject: CWD aka MAD DEER/ELK TO HUMANS ???

Date: September 30, 2002 at 7:06 am PST

From: "Belay, Ermias"

To: Cc: "Race, Richard (NIH)" ; ; "Belay, Ermias"

Sent: Monday, September 30, 2002 9:22 AM

Subject: RE: TO CDC AND NIH - PUB MED- 3 MORE DEATHS - CWD - YOUNG HUNTERS

Dear Sir/Madam,

In the Archives of Neurology you quoted (the abstract of which was attached to your email), we did not say CWD in humans will present like variant CJD.. That assumption would be wrong. I encourage you to read the whole article and call me if you have questions or need more clarification (phone: 404-639-3091). Also, we do not claim that "no-one has ever been infected with prion disease from eating venison." Our conclusion stating that we found no strong evidence of CWD transmission to humans in the article you quoted or in any other forum is limited to the patients we investigated.

Ermias Belay, M.D. Centers for Disease Control and Prevention

-----Original Message-----

From: Sent: Sunday, September 29, 2002 10:15 AM


Subject: TO CDC AND NIH - PUB MED- 3 MORE DEATHS - CWD - YOUNG HUNTERS

Sunday, November 10, 2002 6:26 PM .......snip........end..............TSS

Thursday, April 03, 2008

A prion disease of cervids: Chronic wasting disease 2008 1: Vet Res. 2008 Apr 3;39(4):41 A prion disease of cervids: Chronic wasting disease Sigurdson CJ.

snip...

*** twenty-seven CJD patients who regularly consumed venison were reported to the Surveillance Center***,

snip... full text ; 


> However, to date, no CWD infections have been reported in people. 

sporadic, spontaneous CJD, 85%+ of all human TSE, did not just happen. never in scientific literature has this been proven.

if one looks up the word sporadic or spontaneous at pubmed, you will get a laundry list of disease that are classified in such a way;



key word here is 'reported'. science has shown that CWD in humans will look like sporadic CJD. SO, how can one assume that CWD has not already transmitted to humans? they can't, and it's as simple as that. from all recorded science to date, CWD has already transmitted to humans, and it's being misdiagnosed as sporadic CJD. ...terry 

*** LOOKING FOR CWD IN HUMANS AS nvCJD or as an ATYPICAL CJD, LOOKING IN ALL THE WRONG PLACES $$$ ***

> However, to date, no CWD infections have been reported in people.
key word here is ‘reported’. science has shown that CWD in humans will look like sporadic CJD. SO, how can one assume that CWD has not already transmitted to humans? they can’t, and it’s as simple as that. from all recorded science to date, CWD has already transmitted to humans, and it’s being misdiagnosed as sporadic CJD. …terry
*** LOOKING FOR CWD IN HUMANS AS nvCJD or as an ATYPICAL CJD, LOOKING IN ALL THE WRONG PLACES $$$ ***
*** These results would seem to suggest that CWD does indeed have zoonotic potential, at least as judged by the compatibility of CWD prions and their human PrPC target. Furthermore, extrapolation from this simple in vitro assay suggests that if zoonotic CWD occurred, it would most likely effect those of the PRNP codon 129-MM genotype and that the PrPres type would be similar to that found in the most common subtype of sCJD (MM1).***
CWD TSE PRION AND ZOONOTIC, ZOONOSIS, POTENTIAL

Subject: Re: DEER SPONGIFORM ENCEPHALOPATHY SURVEY & HOUND STUDY 

Date: Fri, 18 Oct 2002 23:12:22 +0100 

From: Steve Dealler 

Reply-To: Bovine Spongiform Encephalopathy Organization: Netscape Online member 

To: BSE-L@ References: <3daf5023 .4080804="" wt.net="">

Dear Terry,

An excellent piece of review as this literature is desparately difficult to get back from Government sites.

What happened with the deer was that an association between deer meat eating and sporadic CJD was found in about 1993. The evidence was not great but did not disappear after several years of asking CJD cases what they had eaten. I think that the work into deer disease largely stopped because it was not helpful to the UK industry...and no specific cases were reported. Well, if you dont look adequately like they are in USA currenly then you wont find any!

Steve Dealler =============== 


''The association between venison eating and risk of CJD shows similar pattern, with regular venison eating associated with a 9 FOLD INCREASE IN RISK OF CJD (p = 0.04).''

CREUTZFELDT JAKOB DISEASE SURVEILLANCE IN THE UNITED KINGDOM THIRD ANNUAL REPORT AUGUST 1994

Consumption of venison and veal was much less widespread among both cases and controls. For both of these meats there was evidence of a trend with increasing frequency of consumption being associated with increasing risk of CJD. (not nvCJD, but sporadic CJD...tss) These associations were largely unchanged when attention was restricted to pairs with data obtained from relatives. ...

Table 9 presents the results of an analysis of these data.

There is STRONG evidence of an association between ‘’regular’’ veal eating and risk of CJD (p = .0.01).

Individuals reported to eat veal on average at least once a year appear to be at 13 TIMES THE RISK of individuals who have never eaten veal.

There is, however, a very wide confidence interval around this estimate. There is no strong evidence that eating veal less than once per year is associated with increased risk of CJD (p = 0.51).

The association between venison eating and risk of CJD shows similar pattern, with regular venison eating associated with a 9 FOLD INCREASE IN RISK OF CJD (p = 0.04).

There is some evidence that risk of CJD INCREASES WITH INCREASING FREQUENCY OF LAMB EATING (p = 0.02).

The evidence for such an association between beef eating and CJD is weaker (p = 0.14). When only controls for whom a relative was interviewed are included, this evidence becomes a little STRONGER (p = 0.08).

snip...

It was found that when veal was included in the model with another exposure, the association between veal and CJD remained statistically significant (p = < 0.05 for all exposures), while the other exposures ceased to be statistically significant (p = > 0.05).

snip...

In conclusion, an analysis of dietary histories revealed statistical associations between various meats/animal products and INCREASED RISK OF CJD. When some account was taken of possible confounding, the association between VEAL EATING AND RISK OF CJD EMERGED AS THE STRONGEST OF THESE ASSOCIATIONS STATISTICALLY. ...

snip...

In the study in the USA, a range of foodstuffs were associated with an increased risk of CJD, including liver consumption which was associated with an apparent SIX-FOLD INCREASE IN THE RISK OF CJD. By comparing the data from 3 studies in relation to this particular dietary factor, the risk of liver consumption became non-significant with an odds ratio of 1.2 (PERSONAL COMMUNICATION, PROFESSOR A. HOFMAN. ERASMUS UNIVERSITY, ROTTERDAM). (???...TSS)

snip...see full report ;




Stephen Dealler is a consultant medical microbiologist  deal@airtime.co.uk 

BSE Inquiry Steve Dealler

Management In Confidence

BSE: Private Submission of Bovine Brain Dealler

snip...see full text;

MONDAY, FEBRUARY 25, 2019

***> MAD DOGS AND ENGLISHMEN BSE, SCRAPIE, CWD, CJD, TSE PRION A REVIEW 2019


***> ''The association between venison eating and risk of CJD shows similar pattern, with regular venison eating associated with a 9 FOLD INCREASE IN RISK OF CJD (p = 0.04).''

***> In conclusion, sensory symptoms and loss of reflexes in Gerstmann-Sträussler-Scheinker syndrome can be explained by neuropathological changes in the spinal cord. We conclude that the sensory symptoms and loss of lower limb reflexes in Gerstmann-Sträussler-Scheinker syndrome is due to pathology in the caudal spinal cord. <***

***> The clinical and pathological presentation in macaques was mostly atypical, with a strong emphasis on spinal cord pathology.<*** 

***> The notion that CWD can be transmitted orally into both new-world and old-world non-human primates asks for a careful reevaluation of the zoonotic risk of CWD. <***

***> All animals have variable signs of prion neuropathology in spinal cords and brains and by supersensitive IHC, reaction was detected in spinal cord segments of all animals.<*** 

***> In particular the US data do not clearly exclude the possibility of human (sporadic or familial) TSE development due to consumption of venison. The Working Group thus recognizes a potential risk to consumers if a TSE would be present in European cervids.'' Scientific opinion on chronic wasting disease (II) <***


North American and Norwegian Chronic Wasting Disease prions exhibit different potential for interspecies transmission and zoonotic risk

Sandra Pritzkow1,*, Damian Gorski1,*, Frank Ramirez1 , Glenn C. Telling2 , Sylvie L. Benestad3 and Claudio Soto1,#

1 Mitchell Center for Alzheimer's disease and related Brain disorders, Department of Neurology, University of Texas McGovern Medical School at Houston, Texas, USA 2 Prion Research Center, Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, Colorado, USA 3 Norwegian Veterinary Institute, OIE Reference Laboratory for CWD, Oslo, Norway.

Summary: We investigated the in vitro spillover and zoonotic potential of CWD from various cervid species. Our results suggest that Norway CWD prions have a higher potential to infect other animals, but NorthAmerican CWD appear more prone to generate human prions.

The current evidence for CWD transmission to humans is controversial; indeed, while transgenic mice expressing human PrP did not develop disease when challenged with CWD prions in various laboratories [6-8, 41], experimental inoculation of CWD into squirrel monkeys produced disease [9, 10]. Studies in macaques, which are phylogenetically closer to humans than squirrel monkeys [45] have shown mixed results. A study from Czub and colleagues found that CWD prions can induce disease and pathologic abnormalities typical of prion disease in macaques exposed to CWD prions, even by oral inoculation of muscle tissue from cervids affected by CWD [46]. However, a different study found no evidence for prion disease in macaques inoculated with CWD [47]. To assess the cervid/human species barrier, we previously used PMCA to determine prion replication in vitro. We found that, after stabilization by successive passages in deer PrPC, PrPSc from CWD infected deer can convert human PrPC into a novel form of PrPSc [13]. Our current study to evaluate in vitro zoonotic potential of various CWD prions showed that although the cervid/human barrier is large, we were able to observe generation of human PrPSc with some specific CWD strains in a second round of PMCA (Fig. 5). The three North American CWD isolates were capable to sustain generation of human PrPSc, with white-tailed deer showing the highest efficiency. Conversely, none of the three Norway CWD isolates generated any detectable PrPSc signal up to the second round of PMCA. This data suggest that North American CWD prions might be of a greater risk to humans than the infected animals in Northern Europe. We speculate that these differences might be due to Norwegian CWD being less stable prion strains as compared to North American CWD, which have had longer time to replicate in cervids and become stabilized through many rounds of natural infection. Our findings may provide important information to understand the diversity of natural CWD prion strains in different animals across distinct geographical areas and their consequences for the spillover into other animal species, including humans.


MONDAY, JULY 19, 2021 

U Calgary researchers at work on a vaccine against a fatal infectious disease affecting deer and potentially people


TUESDAY, JULY 13, 2021

Chronic Wasting Disease and the Canadian Agriculture and Agri-food Sectors Current Knowledge Risks and Policy Options

''The science is progressing on the possibility of transmission of CWD to humans through oral transmission, but the complete assessment of this possibility remains to be done.''


MONDAY, DECEMBER 16, 2019 

Chronic Wasting Disease CWD TSE Prion aka mad cow type disease in cervid Zoonosis Update

***> ''In particular the US data do not clearly exclude the possibility of human (sporadic or familial) TSE development due to consumption of venison. The Working Group thus recognizes a potential risk to consumers if a TSE would be present in European cervids.'' Scientific opinion on chronic wasting disease (II) <***

What if?


TUESDAY, MAY 11, 2021 
 
A Unique Presentation of Creutzfeldt-Jakob Disease in a Patient Consuming Deer Antler Velvet

Conclusion

We believe that our patient’s case of CJD is highly suspicious for cervid etiology given the circumstances of the case as well as the strong evidence of plausibility reported in published literature. This is the first known case of CJD in a patient who had consumed deer antler velvet. Despite the confirmed diagnosis of CJD, a causal relationship between the patient’s disease and his consumption of deer antler velvet cannot be definitively concluded.

Supplemental data including molecular tissue sample analysis and autopsy findings could yield further supporting evidence. Given this patient’s clinical resemblance to CBD and the known histological similarities of CBD with CJD, clinicians should consider both diseases in the differential diagnosis of patients with a similarly esoteric presentation. Regardless of the origin of this patient’s disease, it is clear that the potential for prion transmission from cervids to humans should be further investigated by the academic community with considerable urgency. 


''We believe that our patient’s case of CJD is highly suspicious for cervid etiology given the circumstances of the case as well as the strong evidence of plausibility reported in published literature. This is the first known case of CJD in a patient who had consumed deer antler velvet. Despite the confirmed diagnosis of CJD, a causal relationship between the patient’s disease and his consumption of deer antler velvet cannot be definitively concluded.''
 

ABOUT that deer antler spray and CWD TSE PRION...
 
I have been screaming this since my neighbors mom died from cjd, and she had been taking a supplement that contained bovine brain, bovine eyeball, and other SRMs specified risk materials, the most high risk for mad cow disease.
just saying...
 
I made a submission to the BSE Inquiry long ago during the BSE Inquiry days, and they seemed pretty interested.
 
Sender: "Patricia Cantos"
 
To: "Terry S Singeltary Sr. (E-mail)"
 
Subject: Your submission to the Inquiry
 
Date: Fri, 3 Jul 1998 10:10:05 +0100
 
3 July 1998
 
Mr Terry S Singeltary Sr.
 
E-Mail: Flounder at wt.net
 
Ref: E2979
 
Dear Mr Singeltary,
 
Thank you for your E-mail message of the 30th of June 1998 providing the Inquiry with your further comments.
 
Thank you for offering to provide the Inquiry with any test results on the nutritional supplements your mother was taking before she died.
 
As requested I am sending you our general Information Pack and a copy of the Chairman's letter. Please contact me if your system cannot read the attachments.
 
Regarding your question, the Inquiry is looking into many aspects of the scientific evidence on BSE and nvCJD. I would refer you to the transcripts of evidence we have already heard which are found on our internet site at ;
 
 
Could you please provide the Inquiry with a copy of the press article you refer to in your e-mail? If not an approximate date for the article so that we can locate it?
 
In the meantime, thank you for you comments. Please do not hesitate to contact me on...
 
snip...end...tss
 
everyone I tell this too gets it screwed up...MY MOTHER WAS NOT TAKING THOSE SUPPLEMENTS IPLEX (that I ever knew of). this was my neighbors mother that died exactly one year _previously_ and to the day of sporadic CJD that was diagnosed as Alzheimer’s at first. my mother died exactly a year later from the Heidenhain Variant of Creutzfeldt Jakob Disease hvCJD, and exceedingly rare strains of the ever growing sporadic CJD’s. _both_ cases confirmed. ...kind regards, terry
 
TSEs i.e. mad cow disease's BSE/BASE and NUTRITIONAL SUPPLEMENTS
 
IPLEX, mad by standard process;
 
vacuum dried bovine BRAIN, bone meal, bovine EYE, veal Bone, bovine liver powder, bovine adrenal, vacuum dried bovine kidney, and vacuum dried porcine stomach.
 
also;
 
what about potential mad cow candy bars ?
 
see their potential mad cow candy bar list too...
 
THESE are just a few of MANY of just this ONE COMPANY...TSS
 
DEPARTMENT OF HEALTH AND HUMAN SERVICES
 
FOOD AND DRUG ADMINISTRATION CENTER FOR BIOLOGICS EVALUATION AND RESEARCH
 
TRANSMISSIBLE SPONGIFORM ENCEPHALOPATHIES ADVISORY COMMITTEE
 
Friday, January 19, 2001 snip...
 
17 But I think that we could exhibit some quite
 
18 reasonable concern about blood donors who are taking dietary
 
19 supplements that contain a certain amount of unspecified-
 
20 origin brain, brain-related, brain and pituitary material.
 
21 If they have done this for more than a sniff or something
 
22 like that, then, perhaps, they should be deferred as blood
 
23 donors.
 
24 That is probably worse than spending six months in
 
25 the U.K.
 
1/19/01
 
3681t2.rtf(845) page 501
 
 
 
 
see full text ;
 

Saturday, May 1, 2021 

Clinical Use of Improved Diagnostic Testing for Detection of Prion Disease

 ***Moreover, sporadic disease has never been observed in breeding colonies or primate research laboratories, most notably among hundreds of animals over several decades of study at the National Institutes of Health25, and in nearly twenty older animals continuously housed in our own facility.***

Even if the prevailing view is that sporadic CJD is due to the spontaneous formation of CJD prions, it remains possible that its apparent sporadic nature may, at least in part, result from our limited capacity to identify an environmental origin.

https://www.nature.com/articles/srep11573 

terry