Classical BSE dismissed as the cause of CWD in Norwegian red deer despite strain similarities between both prion agents.

The first case of CWD in a Norwegian red deer was detected by a routine ELISA test and confirmed by western blotting and immunohistochemistry in the brain stem of the animal. Two different western blotting tests were conducted independently in two different laboratories, showing that the red deer glycoprofile was different from the Norwegian CWD reindeer and CWD moose and from North American CWD. The isolate showed nevertheless features similar to the classical BSE (BSE-C) strain. Furthermore, BSE-C could not be excluded based on the PrPSc immunohistochemistry staining in the brainstem and the absence of detectable PrPSc in the lymphoid tissues. Because of the known ability of BSE-C to cross species barriers as well as its zoonotic potential, the CWD red deer isolate was submitted to the EURL Strain Typing Expert Group (STEG) as a BSE-C suspect for further investigation. In addition, different strain typing in vivo and in vitro strategies aiming at identifying the BSE-C strain in the red deer isolate were performed independently in three research groups and BSE-C was not found in it. These results suggest that the Norwegian CWD red deer case was infected with a previously unknown CWD type and further investigation is needed to determine the characteristics of this potential new CWD strain.

Erratum: A tetracationic porphyrin with dual anti-prion activity.

[This corrects the article DOI: 10.1016/j.isci.2023.107480.].

A tetracationic porphyrin with dual anti-prion activity.

Prions are deadly infectious agents made of PrPSc, a misfolded variant of the cellular prion protein (PrPC) which self-propagates by inducing misfolding of native PrPC. PrPSc can adopt different pathogenic conformations (prion strains), which can be resistant to potential drugs, or acquire drug resistance, hampering the development of effective therapies. We identified Zn(II)-BnPyP, a tetracationic porphyrin that binds to distinct domains of native PrPC, eliciting a dual anti-prion effect. Zn(II)-BnPyP binding to a C-terminal pocket destabilizes the native PrPC fold, hindering conversion to PrPSc; Zn(II)-BnPyP binding to the flexible N-terminal tail disrupts N- to C-terminal interactions, triggering PrPC endocytosis and lysosomal degradation, thus reducing the substrate for PrPSc generation. Zn(II)-BnPyP inhibits propagation of different prion strains in vitro, in neuronal cells and organotypic brain cultures. These results identify a PrPC-targeting compound with an unprecedented dual mechanism of action which might be exploited to achieve anti-prion effects without engendering drug resistance.

Strain-Dependent Morphology of Reactive Astrocytes in Human- and Animal-Vole-Adapted Prions.

Reactive astrogliosis is one of the pathological hallmarks of prion diseases. Recent studies highlighted the influence of several factors on the astrocyte phenotype in prion diseases, including the brain region involved, the genotype backgrounds of the host, and the prion strain. Elucidating the influence of prion strains on the astrocyte phenotype may provide crucial insights for developing therapeutic strategies. Here, we investigated the relationship between prion strains and astrocyte phenotype in six human- and animal-vole-adapted strains characterized by distinctive neuropathological features. In particular, we compared astrocyte morphology and astrocyte-associated PrPSc deposition among strains in the same brain region, the mediodorsal thalamic nucleus (MDTN). Astrogliosis was detected to some extent in the MDTN of all analyzed voles. However, we observed variability in the morphological appearance of astrocytes depending on the strain. Astrocytes displayed variability in thickness and length of cellular processes and cellular body size, suggesting strain-specific phenotypes of reactive astrocytes. Remarkably, four out of six strains displayed astrocyte-associated PrPSc deposition, which correlated with the size of astrocytes. Overall, these data show that the heterogeneous reactivity of astrocytes in prion diseases depends at least in part on the infecting prion strains and their specific interaction with astrocytes.

Bona fide atypical scrapie faithfully reproduced for the first time in a rodent model.

Atypical Scrapie, which is not linked to epidemics, is assumed to be an idiopathic spontaneous prion disease in small ruminants. Therefore, its occurrence is unlikely to be controlled through selective breeding or other strategies as it is done for classical scrapie outbreaks. Its spontaneous nature and its sporadic incidence worldwide is reminiscent of the incidence of idiopathic spontaneous prion diseases in humans, which account for more than 85% of the cases in humans. Hence, developing animal models that consistently reproduce this phenomenon of spontaneous PrP misfolding, is of importance to study the pathobiology of idiopathic spontaneous prion disorders. Transgenic mice overexpressing sheep PrPC with I112 polymorphism (TgShI112, 1-2 × PrP levels compared to sheep brain) manifest clinical signs of a spongiform encephalopathy spontaneously as early as 380 days of age. The brains of these animals show the neuropathological hallmarks of prion disease and biochemical analyses of the misfolded prion protein show a ladder-like PrPres pattern with a predominant 7-10 kDa band. Brain homogenates from spontaneously diseased transgenic mice were inoculated in several models to assess their transmissibility and characterize the prion strain generated: TgShI112 (ovine I112 ARQ PrPC), Tg338 (ovine VRQ PrPC), Tg501 (ovine ARQ PrPC), Tg340 (human M129 PrPC), Tg361 (human V129 PrPC), TgVole (bank vole I109 PrPC), bank vole (I109I PrPC), and sheep (AHQ/ARR and AHQ/AHQ churra-tensina breeds). Our analysis of the results of these bioassays concludes that the strain generated in this model is indistinguishable to that causing atypical scrapie (Nor98). Thus, we present the first faithful model for a bona fide, transmissible, ovine, atypical scrapie prion disease.

Gerstmann-Sträussler-Scheinker Disease with F198S Mutation Induces Independent Tau and Prion Protein Pathologies in Bank Voles.

Gerstmann-Sträussler-Scheinker disease (GSS) is a rare genetic prion disease. A large GSS kindred linked to the serine-for-phenylalanine substitution at codon 198 of the prion protein gene (GSS-F198S) is characterized by conspicuous accumulation of prion protein (PrP)-amyloid deposits and neurofibrillary tangles. Recently, we demonstrated the transmissibility of GSS-F198S prions to bank vole carrying isoleucine at 109 PrP codon (BvI). Here we investigated: (i) the transmissibility of GSS-F198S prions to voles carrying methionine at codon 109 (BvM); (ii) the induction of hyperphosphorylated Tau (pTau) in two vole lines, and (iii) compared the phenotype of GSS-F198S-induced pTau with pTau induced in BvM following intracerebral inoculation of a familial Alzheimer's disease case carrying Presenilin 1 mutation (fAD-PS1). We did not detect prion transmission to BvM, despite the high susceptibility of BvI previously observed. Immunohistochemistry established the presence of induced pTau depositions in vole brains that were not affected by prions. Furthermore, the phenotype of pTau deposits in vole brains was similar in GSS-F198S and fAD-PS1. Overall, results suggest that, regardless of the cause of pTau deposition and its relationship with PrPSc in GSS-F198S human-affected brains, the two components possess their own seeding properties, and that pTau deposition is similarly induced by GSS-F198S and fAD-PS1.

A single amino acid residue in bank vole prion protein drives permissiveness to Nor98/atypical scrapie and the emergence of multiple strain variants.

Prions are infectious agents that replicate through the autocatalytic misfolding of the cellular prion protein (PrPC) into infectious aggregates (PrPSc) causing fatal neurodegenerative diseases in humans and animals. Prions exist as strains, which are encoded by conformational variants of PrPSc. The transmissibility of prions depends on the PrPC sequence of the recipient host and on the incoming prion strain, so that some animal prion strains are more contagious than others or are transmissible to new species, including humans. Nor98/atypical scrapie (AS) is a prion disease of sheep and goats reported in several countries worldwide. At variance with classical scrapie (CS), AS is considered poorly contagious and is supposed to be spontaneous in origin. The zoonotic potential of AS, its strain variability and the relationships with the more contagious CS strains remain largely unknown. We characterized AS isolates from sheep and goats by transmission in ovinised transgenic mice (tg338) and in two genetic lines of bank voles, carrying either methionine (BvM) or isoleucine (BvI) at PrP residue 109. All AS isolates induced the same pathological phenotype in tg338 mice, thus proving that they encoded the same strain, irrespective of their geographical origin or source species. In bank voles, we found that the M109I polymorphism dictates the susceptibility to AS. BvI were susceptible and faithfully reproduced the AS strain, while the transmission in BvM was highly inefficient and was characterized by a conformational change towards a CS-like prion strain. Sub-passaging experiments revealed that the main strain component of AS is accompanied by minor CS-like strain components, which can be positively selected during replication in both AS-resistant or AS-susceptible animals. These findings add new clues for a better comprehension of strain selection dynamics in prion infections and have wider implications for understanding the origin of contagious prion strains, such as CS.

Pelodera strongyloides in the critically endangered Apennine brown bear (Ursus arctos marsicanus).

Skin biopsies from 20 Apennine brown bears (Ursus arctos marsicanus), 17 of which displaying skin lesions, were investigated by histopathology. Different degrees of dermatitis characterized by folliculitis and furunculosis accompanied by epidermal hyperplasia and epidermal and follicular hyperkeratosis were detected. In the most severe lesions, the superimposition of traumatic wounds, probably self-induced by scratching, was observed. In 8 out of 17 (47.0%) affected bears, cross- and longitudinally-sectioned nematode larvae were present within the lumen of hair follicles, whose localization and morphological characteristics were consistent with Pelodera strongyloides. P. strongyloides is a free-living saprophytic nematode whose third-stage larvae can invade the skin causing pruritic dermatitis in several mammalian species. This is the first report of Pelodera infection in the brown bear. Although capable of causing primary dermatitis, the finding of Pelodera is not sufficient to conclude that it is the cause of the lesions observed in bears. Nevertheless, the high prevalence of the infection is indicative of a diffuse phenomenon that requires further specific investigations given the interest and conservational relevance of this relict bear population.

Detection and whole genome sequencing of murine norovirus in animal facility in Italy.

Viruses belonging to the genus Norovirus (NoV) of the family Caliciviridae are the major cause of acute viral gastroenteritis worldwide. NoVs are classified into 10 genogroups (GI-GX), and those belonging to the genogroup GV are able to infect several species of rodents. To evaluate the circulation of MNV among mice housed in an Italian facility, sampling was performed over two separate periods, in 2011, and 3 years later in 2014. During the two samplings, 75 fecal samples were collected from healthy mice housed in the animal facility and subjected to RT-PCR for viral detection. After the analysis, 41/75 animals (54.6%) resulted positive for the presence of MNV in feces. Nucleotide sequencing revealed the presence of two MNV variants co-circulating in both 2011 and 2014. One MNV strain was isolated on RAW264.7 cell line, and subjected to full genome sequencing. Our study showed that the murine noroviruses are widespread in the investigated animal facility, despite guidelines for animal care and maintenance. Full genome sequence analysis of the MNV strain described in this study showed a correlation with other strains circulating in Europe. Understanding the molecular epidemiology of this virus should give insight into its natural history and evolution in mice.

Stability of BSE infectivity towards heat treatment even after proteolytic removal of prion protein.

The unconventional infectious agents of transmissible spongiform encephalopathies (TSEs) are prions. Their infectivity co-appears with PrPSc, aberrant depositions of the host's cellular prion protein (PrPC). Successive heat treatment in the presence of detergent and proteolysis by a keratinase from Bacillus licheniformis PWD-1 was shown before to destroy PrPSc from bovine TSE (BSE) and sheep scrapie diseased brain, however data regarding expected reduction of infectivity were still lacking. Therefore, transgenic Tgbov XV mice which are highly BSE susceptible were used to quantify infectivity before and after the bovine brain treatment procedure. Also four immunochemical analyses were applied to compare the levels of PrPSc. After heating at 115 °C with or without subsequent proteolysis, the original BSE infectivity of 106.2-6.4 ID50 g-1 was reduced to a remaining infectivity of 104.6-5.7 ID50 g-1 while strain characteristics were unaltered, even after precipitation with methanol. Surprisingly, PrPSc depletion was 5-800 times higher than the loss of infectivity. Similar treatment was applied on other prion strains, which were CWD1 in bank voles, 263 K scrapie in hamsters and sheep PG127 scrapie in tg338 ovinized mice. In these strains however, infectivity was already destroyed by heat only. These findings show the unusual heat resistance of BSE and support a role for an additional factor in prion formation as suggested elsewhere when producing prions from PrPC. Leftover material in the remaining PrPSc depleted BSE preparation offers a unique substrate for searching additional elements for prion infectivity and improving our concept about the nature of prions.

Sensitive protein misfolding cyclic amplification of sporadic Creutzfeldt-Jakob disease prions is strongly seed and substrate dependent.

Unlike variant Creutzfeldt-Jakob disease prions, sporadic Creutzfeldt-Jakob disease prions have been shown to be difficult to amplify in vitro by protein misfolding cyclic amplification (PMCA). We assessed PMCA of pathological prion protein (PrPTSE) from 14 human sCJD brain samples in 3 substrates: 2 from transgenic mice expressing human prion protein (PrP) with either methionine (M) or valine (V) at position 129, and 1 from bank voles. Brain extracts representing the 5 major clinicopathological sCJD subtypes (MM1/MV1, MM2, MV2, VV1, and VV2) all triggered seeded PrPTSE amplification during serial PMCA with strong seed- and substrate-dependence. Remarkably, bank vole PrP substrate allowed the propagation of all sCJD subtypes with preservation of the initial molecular PrPTSE type. In contrast, PMCA in human PrP substrates was accompanied by a PrPTSE molecular shift during heterologous (M/V129) PMCA reactions, with increased permissiveness of V129 PrP substrate to in vitro sCJD prion amplification compared to M129 PrP substrate. Combining PMCA amplification sensitivities with PrPTSE electrophoretic profiles obtained in the different substrates confirmed the classification of 4 distinct major sCJD prion strains (M1, M2, V1, and V2). Finally, the level of sensitivity required to detect VV2 sCJD prions in cerebrospinal fluid was achieved.

Studies in bank voles reveal strain differences between chronic wasting disease prions from Norway and North America.

Chronic wasting disease (CWD) is a relentless epidemic disorder caused by infectious prions that threatens the survival of cervid populations and raises increasing public health concerns in North America. In Europe, CWD was detected for the first time in wild Norwegian reindeer (Rangifer tarandus) and moose (Alces alces) in 2016. In this study, we aimed at comparing the strain properties of CWD prions derived from different cervid species in Norway and North America. Using a classical strain typing approach involving transmission and adaptation to bank voles (Myodes glareolus), we found that prions causing CWD in Norway induced incubation times, neuropathology, regional deposition of misfolded prion protein aggregates in the brain, and size of their protease-resistant core, different from those that characterize North American CWD. These findings show that CWD prion strains affecting Norwegian cervids are distinct from those found in North America, implying that the highly contagious North American CWD prions are not the proximate cause of the newly discovered Norwegian CWD cases. In addition, Norwegian CWD isolates showed an unexpected strain variability, with reindeer and moose being caused by different CWD strains. Our findings shed light on the origin of emergent European CWD, have significant implications for understanding the nature and the ecology of CWD in Europe, and highlight the need to assess the zoonotic potential of the new CWD strains detected in Europe.

Isolation of infectious, non-fibrillar and oligomeric prions from a genetic prion disease.

Prions are transmissible agents causing lethal neurodegenerative diseases that are composed of aggregates of misfolded cellular prion protein (PrPSc). Despite non-fibrillar oligomers having been proposed as the most infectious prion particles, prions purified from diseased brains usually consist of large and fibrillar PrPSc aggregates, whose protease-resistant core (PrPres) encompasses the whole C-terminus of PrP. In contrast, PrPSc from Gerstmann-Sträussler-Scheinker disease associated with alanine to valine substitution at position 117 (GSS-A117V) is characterized by a small protease-resistant core, which is devoid of the C-terminus. We thus aimed to investigate the role of this unusual PrPSc in terms of infectivity, strain characteristics, and structural features. We found, by titration in bank voles, that the infectivity of GSS-A117V is extremely high (109.3 ID50 U/g) and is resistant to treatment with proteinase K (109.0 ID50 U/g). We then purified the proteinase K-resistant GSS-A117V prions and determined the amount of infectivity and PrPres in the different fractions, alongside the morphological characteristics of purified PrPres aggregates by electron microscopy. Purified pellet fractions from GSS-A117V contained the expected N- and C-terminally cleaved 7 kDa PrPres, although the yield of PrPres was low. We found that this low yield depended on the low density/small size of GSS-A117V PrPres, as it was mainly retained in the last supernatant fraction. All fractions were highly infectious, thus confirming the infectious nature of the 7 kDa PrPres, with infectivity levels that directly correlated with the PrPres amount detected. Finally, electron microscopy analysis of these fractions showed no presence of amyloid fibrils, but only very small and indistinct, non-fibrillar PrPresparticles were detected and confirmed to contain PrP via immunogold labelling. Our study demonstrates that purified aggregates of 7 kDa PrPres, spanning residues ∼90-150, are highly infectious oligomers that encode the biochemical and biological strain features of the original sample. Overall, the autocatalytic behaviour of the prion oligomers reveals their role in the propagation of neurodegeneration in patients with Gerstmann-Sträussler-Scheinker disease and implies that the C-terminus of PrPSc is dispensable for infectivity and strain features for this prion strain, uncovering the central PrP domain as the minimal molecular component able to encode infectious prions. These findings are consistent with the hypothesis that non-fibrillar prion particles are highly efficient propagators of disease and provide new molecular and morphological constraints on the structure of infectious prions.

Cofactor and glycosylation preferences for in vitro prion conversion are predominantly determined by strain conformation.

Prion diseases are caused by the misfolding of a host-encoded glycoprotein, PrPC, into a pathogenic conformer, PrPSc. Infectious prions can exist as different strains, composed of unique conformations of PrPSc that generate strain-specific biological traits, including distinctive patterns of PrPSc accumulation throughout the brain. Prion strains from different animal species display different cofactor and PrPC glycoform preferences to propagate efficiently in vitro, but it is unknown whether these molecular preferences are specified by the amino acid sequence of PrPC substrate or by the conformation of PrPSc seed. To distinguish between these two possibilities, we used bank vole PrPC to propagate both hamster or mouse prions (which have distinct cofactor and glycosylation preferences) with a single, common substrate. We performed reconstituted sPMCA reactions using either (1) phospholipid or RNA cofactor molecules, or (2) di- or un-glycosylated bank vole PrPC substrate. We found that prion strains from either species are capable of propagating efficiently using bank vole PrPC substrates when reactions contained the same PrPC glycoform or cofactor molecule preferred by the PrPSc seed in its host species. Thus, we conclude that it is the conformation of the input PrPSc seed, not the amino acid sequence of the PrPC substrate, that primarily determines species-specific cofactor and glycosylation preferences. These results support the hypothesis that strain-specific patterns of prion neurotropism are generated by selection of differentially distributed cofactors molecules and/or PrPC glycoforms during prion replication.

Characterization of goat prions demonstrates geographical variation of scrapie strains in Europe and reveals the composite nature of prion strains.

Bovine Spongiform Encephalopathy (BSE) is the only animal prion which has been recognized as a zoonotic agent so far. The identification of BSE in two goats raised the need to reliably identify BSE in small ruminants. However, our understanding of scrapie strain diversity in small ruminants remains ill-defined, thus limiting the accuracy of BSE surveillance and spreading fear that BSE might lurk unrecognized in goats. We investigated prion strain diversity in a large panel of European goats by a novel experimental approach that, instead of assessing the neuropathological profile after serial transmissions in a single animal model, was based on the direct interaction of prion isolates with several recipient rodent models expressing small ruminants or heterologous prion proteins. The findings show that the biological properties of scrapie isolates display different patterns of geographical distribution in Europe and suggest that goat BSE could be reliably discriminated from a wide range of biologically and geographically diverse goat prion isolates. Finally, most field prion isolates showed composite strain features, with discrete strain components or sub-strains being present in different proportions in individual goats or tissues. This has important implications for understanding the nature and evolution of scrapie strains and their transmissibility to other species, including humans.

Development of a new largely scalable in vitro prion propagation method for the production of infectious recombinant prions for high resolution structural studies.

The resolution of the three-dimensional structure of infectious prions at the atomic level is pivotal to understand the pathobiology of Transmissible Spongiform Encephalopathies (TSE), but has been long hindered due to certain particularities of these proteinaceous pathogens. Difficulties related to their purification from brain homogenates of disease-affected animals were resolved almost a decade ago by the development of in vitro recombinant prion propagation systems giving rise to highly infectious recombinant prions. However, lack of knowledge about the molecular mechanisms of the misfolding event and the complexity of systems such as the Protein Misfolding Cyclic Amplification (PMCA), have limited generating the large amounts of homogeneous recombinant prion preparations required for high-resolution techniques such as solid state Nuclear Magnetic Resonance (ssNMR) imaging. Herein, we present a novel recombinant prion propagation system based on PMCA that substitutes sonication with shaking thereby allowing the production of unprecedented amounts of multi-labeled, infectious recombinant prions. The use of specific cofactors, such as dextran sulfate, limit the structural heterogeneity of the in vitro propagated prions and makes possible, for the first time, the generation of infectious and likely homogeneous samples in sufficient quantities for studies with high-resolution structural techniques as demonstrated by the preliminary ssNMR spectrum presented here. Overall, we consider that this new method named Protein Misfolding Shaking Amplification (PMSA), opens new avenues to finally elucidate the three-dimensional structure of infectious prions.

Full restoration of specific infectivity and strain properties from pure mammalian prion protein.

The protein-only hypothesis predicts that infectious mammalian prions are composed solely of PrPSc, a misfolded conformer of the normal prion protein, PrPC. However, protein-only PrPSc preparations lack significant levels of prion infectivity, leading to the alternative hypothesis that cofactor molecules are required to form infectious prions. Here, we show that prions with parental strain properties and full specific infectivity can be restored from protein-only PrPSc in vitro. The restoration reaction is rapid, potent, and requires bank vole PrPC substrate, post-translational modifications, and cofactor molecules. To our knowledge, this represents the first report in which the essential properties of an infectious mammalian prion have been restored from pure PrP without adaptation. These findings provide evidence for a unified hypothesis of prion infectivity in which the global structure of protein-only PrPSc accurately stores latent infectious and strain information, but cofactor molecules control a reversible switch that unmasks biological infectivity.

Variable Protease-Sensitive Prionopathy Transmission to Bank Voles.

Variably protease-sensitive prionopathy (VPSPr), a recently described human sporadic prion disease, features a protease-resistant, disease-related prion protein (resPrPD) displaying 5 fragments reminiscent of Gerstmann-Sträussler-Scheinker disease. Experimental VPSPr transmission to human PrP-expressing transgenic mice, although replication of the VPSPr resPrPD profile succeeded, has been incomplete because of second passage failure. We bioassayed VPSPr in bank voles, which are susceptible to human prion strains. Transmission was complete; first-passage attack rates were 5%-35%, and second-passage rates reached 100% and survival times were 50% shorter. We observed 3 distinct phenotypes and resPrPD profiles; 2 imitated sporadic Creutzfeldt-Jakob disease resPrPD, and 1 resembled Gerstmann-Sträussler-Scheinker disease resPrPD. The first 2 phenotypes may be related to the presence of minor PrPD components in VPSPr. Full VPSPr transmission confirms permissiveness of bank voles to human prions and suggests that bank vole PrP may efficiently reveal an underrepresented native strain but does not replicate the complex VPSPr PrPD profile.

Novel Type of Chronic Wasting Disease Detected in Moose (Alces alces), Norway.

Chronic wasting disease (CWD) persists in cervid populations of North America and in 2016 was detected for the first time in Europe in a wild reindeer in Norway. We report the detection of CWD in 3 moose (Alces alces) in Norway, identified through a large scale surveillance program. The cases occurred in 13-14-year-old female moose, and we detected an abnormal form of prion protein (PrPSc) in the brain but not in lymphoid tissues. Immunohistochemistry revealed that the moose shared the same neuropathologic phenotype, characterized by mostly intraneuronal deposition of PrPSc. This pattern differed from that observed in reindeer and has not been previously reported in CWD-infected cervids. Moreover, Western blot revealed a PrPSc type distinguishable from previous CWD cases and from known ruminant prion diseases in Europe, with the possible exception of sheep CH1641. These findings suggest that these cases in moose represent a novel type of CWD.

Prion Disease in Dromedary Camels, Algeria.

Prions cause fatal and transmissible neurodegenerative diseases, including Creutzfeldt-Jakob disease in humans, scrapie in small ruminants, and bovine spongiform encephalopathy (BSE). After the BSE epidemic, and the associated human infections, began in 1996 in the United Kingdom, general concerns have been raised about animal prions. We detected a prion disease in dromedary camels (Camelus dromedarius) in Algeria. Symptoms suggesting prion disease occurred in 3.1% of dromedaries brought for slaughter to the Ouargla abattoir in 2015-2016. We confirmed diagnosis by detecting pathognomonic neurodegeneration and disease-specific prion protein (PrPSc) in brain tissues from 3 symptomatic animals. Prion detection in lymphoid tissues is suggestive of the infectious nature of the disease. PrPSc biochemical characterization showed differences with BSE and scrapie. Our identification of this prion disease in a geographically widespread livestock species requires urgent enforcement of surveillance and assessment of the potential risks to human and animal health.