Article

Review: Update on Classical and Atypical Scrapie in Sheep and Goats

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Abstract

Scrapie is a naturally occurring transmissible spongiform encephalopathy (TSE) or prion disease of sheep and goats. Scrapie is a protein misfolding disease where the normal prion protein (PrP C) misfolds into a pathogenic form (PrP Sc) that is highly resistant to enzymatic breakdown within the cell and accumulates, eventually leading to neurodegeneration. The amino acid sequence of the prion protein and tissue distribution of PrP Sc within affected hosts have a major role in determining susceptibility to and potential environmental contamination with the scrapie agent. Many countries have genotype-based eradication programs that emphasize using rams that express arginine at codon 171 in the prion protein, which is associated with resistance to the classical scrapie agent. In classical scrapie, accumulation of PrP Sc within lymphoid and other tissues facilitates environmental contamination and spread of the disease within flocks. A major distinction can be made between classical scrapie strains that are readily spread within populations of susceptible sheep and goats and atypical (Nor-98) scrapie that has unique molecular and phenotype characteristics and is thought to occur spontaneously in older sheep or goats. This review provides an overview of classical and atypical scrapie with consideration of potential transmission of classical scrapie to other mammalian hosts.

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... During the BSE pandemic, it became evident that prion-contaminated meat and bone meal was fed to cattle, and the resulting infected beef was subsequently consumed by humans [18][19][20]. There is a hypothesis that the prion originated from scrapie, although the origin of scrapie remains unknown [21]. Given that pathogenic single nucleotide polymorphisms (SNPs) have been reported only in humans and that prion seed SNPs have not been reported in non-human prion host animals, prion seeds derived from animals at lower evolutionary stages may be the cause of scrapie. ...
... Research on prion diseases has been conducted on various animals such as sheep, cattle, goats, deer, birds, and dogs, and major concerns of that research are the susceptibility and resistance to prion diseases according to genetic polymorphisms in the PRNP gene [5,12,21,[43][44][45]. In previous studies, the risk of classical scrapie can be classified into five types according to the 136 (A>V), 154 (R>H), and 171 (R>Q, H) polymorphisms in the ovine PRNP gene in sheep [21]. ...
... Research on prion diseases has been conducted on various animals such as sheep, cattle, goats, deer, birds, and dogs, and major concerns of that research are the susceptibility and resistance to prion diseases according to genetic polymorphisms in the PRNP gene [5,12,21,[43][44][45]. In previous studies, the risk of classical scrapie can be classified into five types according to the 136 (A>V), 154 (R>H), and 171 (R>Q, H) polymorphisms in the ovine PRNP gene in sheep [21]. In addition, in the PRNP gene of cattle, a 12 bp deletion in the promoter was associated with BSE susceptibility, and an additional 23 bp deletion in the same promoter showed the highest susceptibility to BSE [12]. ...
Article
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Prion diseases are fatal neurodegenerative diseases that can be transmitted by infectious protein particles, PrPScs, encoded by the endogenous prion protein gene (PRNP). The origin of prion seeds is unclear, especially in non-human hosts, and this identification is pivotal to preventing the spread of prion diseases from host animals. Recently, an abnormally high amyloid propensity in prion proteins (PrPs) was found in a frog, of which the genetic variations in the PRNP gene have not been investigated. In this study, genetic polymorphisms in the PRNP gene were investigated in 194 Dybowski’s frogs using polymerase chain reaction (PCR) and amplicon sequencing. We carried out in silico analyses to predict functional alterations according to non-synonymous single nucleotide polymorphisms (SNPs) using PolyPhen-2, PANTHER, SIFT, and MutPred2. We used ClustalW2 and MEGA X to compare frog PRNP and PrP sequences with those of prion-related animals. To evaluate the impact of the SNPs on protein aggregation propensity and 3D structure, we utilized AMYCO and ColabFold. We identified 34 novel genetic polymorphisms including 6 non-synonymous SNPs in the frog PRNP gene. The hydrogen bond length varied at codons 143 and 207 according to non-synonymous SNPs, even if the electrostatic potential was not changed. In silico analysis predicted S143N to increase the aggregation propensity, and W6L, C8Y, R211W, and L241F had damaging effects on frog PrPs. The PRNP and PrP sequences of frogs showed low homology with those of prion-related mammals. To the best of our knowledge, this study was the first to discover genetic polymorphisms in the PRNP gene in amphibians.
... Atypical scrapie (AS), first detected in 1998 [8], differs from CS in clinical signs, the distribution of PrP Sc and the prion protein genotypes affected [9,10]. In contrast to CS, where within-flock clustering is evident [11], AS cases occur sporadically; cases usually occur as single events, and it is rare for a second case to occur in a given holding [12,13]. ...
... It was therefore concluded that AS is more likely to be non-contagious [1]. As such, AS is recognized as a nonreportable disease separate to CS by the World Organization for Animal Health [9]. ...
... The drawback of using the NSP genotype groupings is that it potentially includes relatively high-risk and low-risk genotypes in the same group, making it difficult to infer the risk by individual genotype. It has been shown that the alleles most associated with AS infection are AHQ and ARQ, [16,22,25,26] which are Group III according to the NSP genotype groupings, with the AS risk of the latter genotype additionally being increased by a polymorphism at codon 141 (F, phenylalanine) [9,25] to give the AFRQ allele. It would be possible to re-run the back-calculation model to estimate AS risks at the individual genotype level, although it is unclear whether the model would be able to produce results with sufficient certainty for the whole range of AS susceptible genotypes, as there would likely be too few samples for some of the genotypes. ...
Article
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Atypical scrapie (AS) is a transmissible spongiform encephalopathy (TSE) that affects sheep and goats. Low within-flock incidence suggests that AS is not transmissible between animals, and testing of all animals that exit positive flocks for two years following detection (i.e., intensified monitoring) used to be carried out in the EU to provide data to test this. This intensified monitoring stopped in 2021 but continues in Great Britain (GB). The aim of this study was to predict the number of AS cases missed if this monitoring were also stopped in GB, using a combination of statistical and transmission modelling. The number of AS cases estimated to be missed if the intensified monitoring was stopped was low relative to the number of AS cases detected in other active surveillance streams (e.g., fallen stock and abattoir surveys), at approximately 1 case every 3 years (0.34 per year, 95% CI: 0.18–0.54) compared to 10 per year (95% CI: 4–17) in the active surveillance stream. This suggests that stopping the intensive monitoring of AS would have relatively little impact on AS surveillance and on the power of the available AS data to infer whether AS is contagious.
... TSEs are fatal neurodegenerative disorders caused by prionsdisease-associated misfolded proteins that replicate their isoforms using a template-like mechanism [1][2][3]. Scrapie and chronic wasting disease (CWD) affecting cervids are contagious among susceptible animals [4][5][6]. Two major classifications of scrapie are recognized: classical scrapie and atypical (Nor98) scrapie. Classical scrapie presents with hind limb ataxia and/or weakness, head tremors, behavioural changes, abnormal posture and gait, weight loss, and pruritus (skin itching) which leads to sheep rubbing against objects and losing their wool [7,p.60-71, ...
... 8-11]. Atypical scrapie is usually detected prior to onset of clinical disease by prion surveillance programmes; clinical signs are typically characterized by ataxia (often in the hind limbs), behavioural changes, and weight loss, in the absence pruritus [6,12,13]. ...
... One explanation is that the Wilkshire and Dorsetshire cases represent a strain of scrapie resembling atypical Nor98 where pruritus is absent and hind limb ataxia can be present (although hind limb ataxia can also be present in classical scrapie) [10,13]. Nor98 is, however, poorly transmitted [6,13,24], whereas Lisle noted outbreaks. Apparent outbreaks of a Nor98-like disease could be explained by the extreme inbreeding of the Wiltshire Horn and Dorset Horn sheep conferring a high degree of genetic susceptibility to scrapie [7,p.8-11, 10, 13]. ...
Article
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Eighteenth-century England witnessed the emergence of two neurological diseases in animals. Scrapie, a transmissible spongiform encephalopathy, is a fatal neurodegenerative disease of sheep and goats that appears in classical and atypical forms. Reports of classical scrapie in continental Europe with described symptoms date back to 1750 in what is now western Poland. However, two major outbreaks of scrapie appeared in England prior to the 1800s. References to a sheep disease with a resemblance to scrapie first appear in Southwestern England between 1693 and 1722 and in the East Midlands between 1693 and 1706. Concurrent with the descriptions of scrapie in sheep was a neurological disease of deer first appearing in the East of England. Two 18th-century writers remarked on the symptomatic similarities between the sheep and deer neurological diseases. Multiple outbreaks of the unknown deer disease existing as early as 1772 are examined and are identified as rabies.
... Der Name kommt von "to scrape", weil erkrankte Schafe aufgrund des starken Juckreizes dazu tendieren, sich an Zaunpfählen oder Toren zu kratzen und zu schrubben. Darüber hinaus manifestiert sich Scrapie bei Schafen, ähnlich wie die Creutzfeldt-Jakob-Erkrankung bei Menschen in bizarren Verhaltensweisen, Schwierigkeiten mit dem Gehen, der Koordination sowie der Balance (Greenlee, 2019;Zetterberg, 2019). ...
... Es zeigten sich progressive Schlaflosigkeit, Dysautonomia (Entgleisung des peripheren und autonomen Nervensystems), Muskelzuckungen und in manchen Fällen auch Demenz (Collins, McLean und Masters, 2001;Llorens u. a., 2017). Nach einer Krankheitszeit von 13 bis 15 Monaten endet sie tödlich (Llorens u. a., 2017 (Wells u. a., 1987 (Greenlee, 2019;Zetterberg, 2019). ...
... In unterschiedlichen Ländern haben sich verschiedene Bezeichnungen für diese Erkrankung entwickelt. Im Englischen heißt sie Scrapie von "to scrape" gleich kratzen, in Deutschland Traberkrankheit wegen des veränderten Gangs der erkrankten Tiere (Hörnlimann, Beat, Detlev Riesner, 2001;Ulvund, 2001 (Foote u. a., 1993;Onodera u. a., 1993;Greenlee, 2019;Cassmann und Greenlee, 2020). Bedingt durch die hohe Stabilität des Erregers können sich so adulte Tiere durch kontaminiertes Weidegras direkt und indirekt infizieren. ...
Thesis
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Die Erreger der TSE, die sogenannten Prionen sind infektiöse Proteine, die für eine Reihe von tödlich verlaufenden neurodegenerativen Erkrankungen beim Menschen und bei Tieren verantwortlich sind. In der vorliegenden Arbeit wurde im Rahmen des Projekts “PRIORITY 222887 Protecting the food chain from prions: shaping European priorities through basic and applied research” Untersuchungen zur Inaktivierung von mit Scrapie 263 K und BSE 301 V versetztem Fleischbrei der Kategorie 1 und 2 durchgeführt. Zum Material der Kategorie 1 gehört spezifische Risikomaterial, welches bei der Schlachtung anfällt und ein potentielles TSE-Risiko birgt. Dieses Material muss zurzeit laut den gesetzlichen Bestimmungen in Europa sterilisiert und verbrannt werden und kann somit nicht einer weiteren Nutzung zugeführt werden. Es sollte eine Methode gefunden werden, mit der zum einen Prion-Proteine sicher inaktiviert werden und zum Anderen eine weitere stoffliche Nutzung dieses hygienisierten Materials ermöglicht wird. Dieses Material könnte z.B. als Phosphor-, Aminosäurequelle in Düngemitteln oder als Teil erneuerbarer Energien wie Biogas oder Biodiesel erfolgen. Die Behandlung mit der Thermodruckhydrolyse mit Mikrowellen ist bereits bei einer Temperatur von 120°C und einer Behandlungsdauer von 5 min erfolgreich in der Inaktivierung des mit Scrapie 263 K versetzten Fleischbrei der Kategorie 1 und 2. Für eine vollständige Inaktivierung des mit BSE 301 V versetzten Fleischbreis der Kategorie 1 und 2 benötigt man eine Temperatur von 120°C und eine Behandlungsdauer von 10 min. Der Nachweis erfolgte hier ausschließlich im Western Blotting. Da der Nachweis im Western Blotting allein nicht belastbar ist, sollte ein Bioassay erfolgen. Dieser war aus verschiedenen Gründen nicht gewünscht, weshalb nach einer Alternative gesucht wurde. Mit Hilfe der in vitro-Amplifizierung, der sogenannte Protein Misfolding Cyclic Amplification (PMCA) wurde eine Methode gefunden, die den Bioassay ersetzen kann. Durch die PMCA lassen sich auch geringe Mengen an Prionen identisch so vervielfältigen, dass diese mittels Western Blotting nachweisbar sind. In den nachfolgenden Inaktivierungsversuchen von mit Scrapie 263 K versetztem Peptonwasser, welches als Fleischbreiersatz diente, wurde bei der Behandlung in der Thermodruckhydrolyse mit Mikrowellen bei einer Behandlungstemperatur von 120°C und 15 min eine vollständige Inaktivierung (mindestens 4-5 log10-Stufen Reduktion) erreicht. Zusammenfassend konnten die Ergebnisse der Arbeit zeigen, dass die Thermodruckhydrolyse mit Mikrowellen geeignet ist, eine verlässliche Inaktivierung von Prion-Proteinen in tierischen Nebenprodukten zu gewährleisten.
... Often, disease phenotypes within a given species are denoted as different 'strains' [24]. Strains in a natural host are commonly defined by the molecular weight profile of the PrP Sc [25], genotypes of susceptible hosts [23], age of disease onset [26,27], and distribution and intensity of misfolded PrP Sc [20,28]. Because the molecular profile on a Western blot may be different depending on host species or strain of infection, to follow is a brief discussion of the significance. ...
... Following the first description of atypical (Nor-98) scrapie in 2003, discussion of scrapie strains is typically framed using classical and atypical scrapie in sheep [3]. There are differences in the molecular weight profile between classical and atypical strains of scrapie, as atypical scrapie has a smaller PrP Sc fragment, is more PK sensitive, and has 5 less intense Western blot bands compared to the three strong bands in classical scrapie [3,4,26]. In sheep, classical scrapie is spread primarily via horizontal transmission shortly after animals are born, though the average age of affected animals is 2-5 years of age [55], and there is widespread distribution of PrP Sc in lymphoid tissues [56,57]. ...
... Atypical scrapie in sheep appears to be sporadic, in that new cases appear in isolation, as opposed to a cluster of infected animals [3], and there is little to no accumulation of PrP Sc in the lymphoid system [4,58]. There are a number of polymorphisms in the PRNP gene of sheep that affect susceptibility and incubation time of different strains of scrapie (reviewed in [26,59]). Experimental and epidemiological evidence suggests that there is an extremely low likelihood that scrapie could be transmitted to humans [29,60]. ...
Article
Full-text available
Prion diseases, also known as transmissible spongiform encephalopathies (TSEs), are a group of neurodegenerative protein misfolding diseases that invariably cause death. TSEs occur when the endogenous cellular prion protein (PrPC) misfolds to form the pathological prion protein (PrPSc), which templates further conversion of PrPC to PrPSc, accumulates, and initiates a cascade of pathologic processes in cells and tissues. Different strains of prion disease within a species are thought to arise from the differential misfolding of the prion protein and have different clinical phenotypes. Different strains of prion disease may also result in differential accumulation of PrPSc in brain regions and tissues of natural hosts. Here, we review differential accumulation that occurs in the retinal ganglion cells, cerebellar cortex and white matter, and plexuses of the enteric nervous system in cattle with bovine spongiform encephalopathy, sheep and goats with scrapie, cervids with chronic wasting disease, and humans with prion diseases. By characterizing TSEs in their natural host, we can better understand the pathogenesis of different prion strains. This information is valuable in the pursuit of evaluating and discovering potential biomarkers and therapeutics for prion diseases.
... The first reported case of CS occurred in the United Kingdom in 1732. This disease was transmitted further through the export of live animals to western European countries, eventually reaching a worldwide distribution, except for Australia and New Zealand [5,6]. CS can result in serious economic loss due to decreased production, export loss, and increased cost for carcass disposal [5]. ...
... This disease was transmitted further through the export of live animals to western European countries, eventually reaching a worldwide distribution, except for Australia and New Zealand [5,6]. CS can result in serious economic loss due to decreased production, export loss, and increased cost for carcass disposal [5]. ...
... Global distribution of AS exhibiting similar incidence and epidemiology within a given flock suggests AS has a spontaneous etiology or is a poorly transmitted TSE under natural conditions. Thus, in contrast to CS, AS is considered a non-reportable disease by the World Organization for Animal Health (OIE) [5,7,13]. ...
Article
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Wasting disease in small ruminants is frequently detected at slaughterhouses. The wasting disorder is manifested by the deterioration of the nutritional and physiological state of the animal indicated by thinness, emaciation, and cachexia. Evidence of emaciation and cachexia, alone, are pathological conditions leading to carcass condemnation during an inspection. Several diseases are associated with a wasting condition, including scrapie, pseudotuberculosis, tuberculosis, paratuberculosis, Maedi Visna, and tumor diseases. On the other hand, parasitic diseases, nutrition disorders, exposure or ingestion of toxins, metabolic conditions, inadequate nutrition due to poor teeth, or poor alimentary diet are conditions contributing to poor body condition. Classical and atypical scrapie is naturally occurring transmissible spongiform encephalopathies in small ruminants. The etiological agent for each one is prions. However, each of these scrapie types is epidemiologically, pathologically, and biochemically different. Though atypical scrapie occurs at low incidence, it is consistently prevalent in the small ruminant population. Hence, it is advisable to include differential diagnosis of this disease, from other possibilities, as a cause of wasting conditions detected during meat inspection at the abattoir. This manuscript is a review of the measures in force at the abattoir for scrapie control, focusing on the differential diagnosis of gross lesions related to wasting conditions detected in small ruminants during meat inspection.
... After oral exposure, PrPSc accumulates in the gut-associated lymphoid tissues (GALT) and can be detected in the tonsils, spleen, and retropharyngeal and mesenteric lymph nodes for months before reaching CNS [7]. During that time, infected sheep can shed the agent into the environment via body fl uids including placental fl uids, milk and colostrum resulting in vertical transmission to newborns, as well as horizontal transmission within the fl ock at the time of lambing [8]. ...
... Three major sites in prnp associated with sheep resistance to classic scrapie are at codons 136, 154, and 171 [9]. Each allele is marked with a three-letter code and the ARR allele is associated with a highly protective effect against infection with classical scrapie but does not provide resistance towards atypical scrapie [8]. Atypical scrapie is not an infectious disease and appears sporadically as a degenerative brain disease in elderly sheep [6]. ...
... Atypical scrapie is not an infectious disease and appears sporadically as a degenerative brain disease in elderly sheep [6]. Susceptibility to classical scrapie is associated with valine (V), arginine (R) and glutamine (Q) on codons 136, 154 and 171 respectively, while alanine (A), histidine (H) and arginine (R) on the same codons provide resistance [8,10,11]. The most common haplotypes are ARR, ARQ, AHQ, ARH and VRQ [12]. ...
Article
Full-text available
Scrapie is an infectious neurodegenerative disease affecting the central nervous system of sheep and goats that belongs to transmissible spongiform encephalopathies. The disease is caused by the accumulation of proteinase-resistant isoform of the prion protein. The sheep predisposition to scrapie is associated with polymorphisms of the PrP gene. Genetic susceptibility to scrapie is mainly related to codons 136, 154, and 171. ARR sheep are strongly scrapie resistant and VRQ genotype is the most susceptible. Many countries have scrapie eradication programs based on using rams with resistant genotype. The eradication program has not yet been implemented in the Republic of Serbia. To examine the genetic makeup of sheep in Serbia related to scrapie, we optimized TaqMan probes of real-time polymerase chain reaction (qPCR) technique for three codons. Blood samples from 100 sheep were analyzed by qPCR and the majority of the examined sheep were AA homozygous for the 136 codon. For codon 154 the most frequent genotype was RR and for codon 171 the most frequent genotype was QQ.
... Figure 3 presents the missense mutations detected in PrP protein from Ovis aries. Variants are classified above and below the bar as 'Risk factor' and 'Resistance' according to in vivo and in vitro analyses [11][12][13][14]. The other variants were analysed by an in-silico approach through the PROVEAN algorithm (http://provean.jcvi.org/index.php, ...
... accessed on 27 January 2021) and classified as "Deleterious" and "Neutral". All the missense mutations were gathered from Ensembl (EN-SOART00020017839.1) and through database searching [13][14][15][16][17][18]. The alignment was performed using T-Coffee program [9] and edited using Genedoc software [10]. ...
... Figure 3 presents the missense mutations detected in PrP protein from Ovis aries. Variants are classified above and below the bar as 'Risk factor' and 'Resistance' according to in vivo and in vitro analyses [11][12][13][14]. The other variants were analysed by an in-silico approach through the PROVEAN algorithm (http://provean.jcvi. ...
Article
Full-text available
Transmissible Spongiform Encephalopathies (TSEs) or prion diseases are a fatal group of infectious, inherited and spontaneous neurodegenerative diseases affecting human and animals. They are caused by the conversion of cellular prion protein (PrPC) into a misfolded pathological isoform (PrPSc or prion- proteinaceous infectious particle) that self-propagates by conformational conversion of PrPC. Yet by an unknown mechanism, PrPC can fold into different PrPSc conformers that may result in different prion strains that display specific disease phenotype (incubation time, clinical signs and lesion profile). Although the pathways for neurodegeneration as well as the involvement of brain inflammation in these diseases are not well understood, the spongiform changes, neuronal loss, gliosis and accumulation of PrPSc are the characteristic neuropathological lesions. Scrapie affecting small ruminants was the first identified TSE and has been considered the archetype of prion diseases, though atypical and new animal prion diseases continue to emerge highlighting the importance to investigate the lesion profile in naturally affected animals. In this report, we review the neuropathology and the neuroinflammation of animal prion diseases in natural hosts from scrapie, going through the zoonotic bovine spongiform encephalopathy (BSE), the chronic wasting disease (CWD) to the newly identified camel prion disease (CPD). CITATION: Orge, L.; Lima, C.; Machado, C.; Tavares, P.; Mendonça, P.; Carvalho, P.; Silva, J.; Pinto, M.d.L.; Bastos, E.; Pereira, J.C.; Gonçalves-Anjo, N.; Gama, A.; Esteves, A.; Alves, A.; Matos, A.C.; Seixas, F.; Silva, F.; Pires, I.; Figueira, L.; Vieira-Pinto, M.; Sargo, R.; Pires, M.d.A. Neuropathology of Animal Prion Diseases. Biomolecules 2021, 11, 466. https://doi.org/10.3390/biom11030466
... Figure 3 presents the missense mutations detected in PrP protein from Ovis aries. Variants are classified above and below the bar as 'Risk factor' and 'Resistance' according to in vivo and in vitro analyses [11][12][13][14]. The other variants were analysed by an in-silico approach through the PROVEAN algorithm (http://provean.jcvi.org/index.php, ...
... accessed on 27 January 2021) and classified as "Deleterious" and "Neutral". All the missense mutations were gathered from Ensembl (EN-SOART00020017839.1) and through database searching [13][14][15][16][17][18]. The alignment was performed using T-Coffee program [9] and edited using Genedoc software [10]. ...
... Figure 3 presents the missense mutations detected in PrP protein from Ovis aries. Variants are classified above and below the bar as 'Risk factor' and 'Resistance' according to in vivo and in vitro analyses [11][12][13][14]. The other variants were analysed by an in-silico approach through the PROVEAN algorithm (http://provean.jcvi. ...
Article
Full-text available
Transmissible Spongiform Encephalopathies (TSEs) or prion diseases are a fatal group of infectious, inherited and spontaneous neurodegenerative diseases affecting human and animals. They are caused by the conversion of cellular prion protein (PrPC) into a misfolded pathological isoform (PrPSc or prion- proteinaceous infectious particle) that self-propagates by conformational conversion of PrPC. Yet by an unknown mechanism, PrPC can fold into different PrPSc conformers that may result in different prion strains that display specific disease phenotype (incubation time, clinical signs and lesion profile). Although the pathways for neurodegeneration as well as the involvement of brain inflammation in these diseases are not well understood, the spongiform changes, neuronal loss, gliosis and accumulation of PrPSc are the characteristic neuropathological lesions. Scrapie affecting small ruminants was the first identified TSE and has been considered the archetype of prion diseases, though atypical and new animal prion diseases continue to emerge highlighting the importance to investigate the lesion profile in naturally affected animals. In this report, we review the neuropathology and the neuroinflammation of animal prion diseases in natural hosts from scrapie, going through the zoonotic bovine spongiform encephalopathy (BSE), the chronic wasting disease (CWD) to the newly identified camel prion disease (CPD).
... In sheep, three polymorphic codons of the PRNP (codons 136, 154, and 171) give rise to five main different alleles, namely VRQ, ARQ, AHQ, ARH, and ARR (defined with the IUPAC amino acid codes). The VRQ/VRQ, VRQ/ARQ, and ARQ/ARQ genotypes are associated with high susceptibility to classical scrapie [12], while the ARR-containing genotypes, with the exception of VRQ/ARR, have been associated with various levels of resistance to disease [13]. On this basis, according to European Union Decision 2003/100/EC (EU 2003) [14], a selection breeding program to increase the frequency of the resistanceassociated ARR allele in sheep populations has been adopted by each member state in the European Community. ...
... Many European studies showed that allele variation and polymorphisms of the PRNP gene can determine the susceptibility or resistance to classical scrapie in goats, but only a few of them have been proven to be associated with resistance [18]. In particular, at least 50 PRNP polymorphisms have been described in goat breeds [19,20], but their involvement in the modulation of the incubation times and in susceptibility to classical scrapie is less defined than in sheep [13,21]. Furthermore, another PRNP variant that contains three repeats of an octapeptide instead of the usual five repeats was found by Goldman [22]. ...
... In particular, this allele is present in cosmopolitan breeds with a low percentage (9.26% in AL and 2.63% in SA), but it is rather frequent in the Italian native goats with a range between 4.17% (BM) and 12.50% (CP, GA). Further alleles with a frequency equal or higher of 5% are moreover observed in BM (4), CP (5,6,10,14,19), GC (5,14), GM (13,22,23), AL (4,8,17), and SA (4,8,18). Regarding the alleles common to several populations/breeds, the following have to be considered: allele 5 (BM, CP, FV, FL, GC, TE) and allele 14 (BM, CP, FL, GC, TE). ...
Article
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In goats, as in sheep, genotypes of the prion protein gene (PRNP) can influence animals’ susceptibility to scrapie. Since the polymorphic codons in sheep are well known, a genetic selection plan has been implemented in Europe, in order to reduce the prevalence of susceptible genotypes to scrapie. In Italy, no breeding plan for scrapie resistance in goats has been adopted, yet. Likewise, according to the most recent modification of Regulation EU 999/2001 (Regulation EU 772/2020) of the European Commission (EU), based on all the available experimental and in field data, K222, D146 and S146 polymorphisms could be used as scrapie resistance alleles in genetic management both in scrapie outbreaks and in disease prevention. In order to collect data on the variability of PRNP, the present study aimed to analyze the sequence of the PRNP gene in eight Italian local goat populations/breeds reared in central and southern Italy (Bianca Monticellana, Capestrina, Facciuta della Valnerina, Fulva del Lazio, Garganica, Grigia Ciociara, Grigia Molisana, and Teramana), some of which were investigated for the first time; moreover, two cosmopolitan breeds (Alpine and Saanen) were included. Blood samples were collected from 219 goats. Genomic DNA was extracted from whole blood. DNA was used as template in PCR amplification of the entire PRNP open reading frame (ORF). Purified amplicons have been sequenced and aligned to Capra hircusPRNP. Particularly, the alleles carrying the resistance-related 222 K polymorphism occurred in all populations with a frequency between 2.5% and 12.5%. An additional resistance allele carrying the S146 variant was observed with a frequency of 3.7% only in the Alpine breed. For three of the estimated alleles, we could not establish if the found double polymorphisms in heterozygosis were in phase, due to technical limitations. In this context, in addition to selective culling in scrapie outbreaks according to the European regulation in force, in the future, selection plans could be adopted to deal with scrapie and to control its diffusion, meanwhile paying attention to preserve a high variability of PRNP.
... The susceptibility of animals to scrapie is predominantly controlled by polymorphism in the PRNP gene [17]. Based on this, different single nucleotide mutations have been identified at codons 136 (A > V), 154 (R > H), and 171 (R > Q/H) of PRNP gene [18,19]. Amino acids at codons 141 and 154 have been found to be associated with various forms of classical scrapie through modification of the configuration of prion protein [20,21]. ...
... Amino acids at codons 141 and 154 have been found to be associated with various forms of classical scrapie through modification of the configuration of prion protein [20,21]. Changes at codon 136 from A to V has been reported to increase susceptibility to scrapie while variations at codon 171 from Q to R cause resistance in sheep [18,22]. ...
Article
Full-text available
Polymorphism of the prion protein gene (PRNP) gene determines an animal’s susceptibility to scrapie. Three polymorphisms at codons 136, 154, and 171 have been linked to classical scrapie susceptibility, although many variants of PRNP have been reported. However, no study has investigated scrapie susceptibility in Nigerian sheep from the drier agro-climate zones. In this study, we aimed to identify PRNP polymorphism in nucleotide sequences of 126 Nigerian sheep by comparing them with public available studies on scrapie-affected sheep. Further, we deployed Polyphen-2, PROVEAN, and AMYCO analyses to determine the structure changes produced by the non-synonymous SNPs. Nineteen (19) SNPs were found in Nigerian sheep with 14 being non-synonymous. Interestingly, one novel SNP (T718C) was identified. There was a significant difference (P < 0.05) in the allele frequencies of PRNP codon 154 between sheep in Italy and Nigeria. Based on the prediction by Polyphen-2, R154H was probably damaging while H171Q was benign. Contrarily, all SNPs were neutral via PROVEAN analysis while two haplotypes (HYKK and HDKK) had similar amyloid propensity of PRNP with resistance haplotype in Nigerian sheep. Our study provides valuable information that could be possibly adopted in programs targeted at breeding for scrapie resistance in sheep from tropical regions.
... Prion diseases are progressive, ultimately fatal, and have no known cure [2]. Scrapie, the TSE of sheep and goats, is broadly categorized into two forms, classical and atypical [3][4][5]. The term atypical scrapie has also been used to refer to prion disease in sheep and goats arising from non-native prions such as bovine spongiform encephalopathy (BSE) and chronic wasting disease (CWD). ...
... During atypical scrapie, PrP Sc accumulates in the brain and this tissue is infectious to sheep through experimental inoculation by intracranial [17][18][19][20][21] and oral [22] routes. PrP Sc accumulation in peripheral tissues, however, has not been detected by immunoassay [3,5,23], which may explain the lack of significant evidence for natural transmission [24]. However, infectivity from peripheral tissues has been documented in the context of transgenic mouse bioassays [19], raising again a concern for potential sources of infective material for natural transmission. ...
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Nor98-like atypical scrapie is a sporadic disease that affects the central nervous system of sheep and goats that, in contrast to classical scrapie, is not generally regarded as naturally transmissible. However, infectivity has been demonstrated via bioassay not only of brain tissue but also of certain peripheral nerves, lymphoid tissues, and muscle. This study examines placental tissue, a well characterized route of natural transmission for classical scrapie. Further, this study was conducted in sheep homozygous for the classical scrapie resistant ARR genotype and is the first to characterize the transmission of Nor98-like scrapie between homozygous-ARR sheep. Nor98-like scrapie isolated from a United States ARR/ARR sheep was transmitted to four ARR/ARR ewes via intracerebral inoculation of brain homogenate. These ewes were followed and observed to 8 years of age, remained non-clinical but exhibited progression of infection that was consistent with Nor98-like scrapie, including characteristic patterns of PrPSc accumulation in the brain and a lack of accumulation in peripheral lymphoid tissues as detected by conventional methods. Immunoblots of placental tissues from the infected ewes revealed accumulation of a distinct conformation of PrPres, particularly as the animals aged; however, the placenta showed no infectivity when analyzed via ovinized mouse bioassay. Taken together, these results support a low risk for natural transmission of Nor98-like scrapie in ARR/ARR sheep.
... Scrapie was first reported in U.S. sheep and goats in 1947 and 1969, respectively, and has since been considered a major health issue in both small ruminants. The presence of scrapie in the U.S. costs around $20 million annually due to significant production losses, export loss of sheep and goat products including genetic materials, and increased expenditures for carcass disposal [9]. ...
... The scrapie eradication program in sheep demonstrated that it is possible to substantially mitigate the prevalence of scrapie using PRNP genotype variabilities [39]; however, resistant alleles were present in over 60% of the US sheep population in the early period of genetic testing for sheep [9]. Despite the low incidence of scrapie in goat population [40], goats may still serve as a scrapie reservoir; therefore, determining the appropriate candidate genotypes present in the goat population will allow for breeding programs that promote scrapie resistance. ...
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Scrapie is a slowly progressive neurodegenerative disease of small ruminants caused by an accumulation of an abnormal isoform of prion protein in the central nervous system. Polymorphisms of the prion protein gene (PRNP) strongly modulate scrapie resistance and incubation period in goats. The aim of this study was to identify PRNP genetic variability in goats across the United States. Blood from a total of 6,029 apparent scrapie disease-free goats from 654 operations and 19 breeds were analyzed. Sequencing of PRNP revealed 26 genotypes with different rates based on eight codons. The GG127, RR154, and QQ222 genotypes were predominant and showed a remarkably high rate across all goats. The QK222 and NS146 genotypes, known to be protective against scrapie, were found in 0.6% [with 95% CI = (0.3, 1.2)] and 22.0% [95% CI = (19.1, 25.2)] of goats, respectively. The QK222 genotype was found in 23.1% of Oberhasli goats tested, with 95%CI = (3.9, 68.7)] and 22.0% of Toggenburg goats tested with 95%CI = (9.7, 42.5)], while NS146 was found in 65.5% of Savannah goats tested, with 95%CI = (30.8, 89.9), 36.7% of Boer goats tested, with 95%CI = (33.1, 40.4), 36.3% of Nubian goats tested, with 95%CI = (27.0, 46.7)], and 35.6% of LaMancha goats tested, with 95%CI = (22.8, 50.8%). The MM142 and IM142 genotypes were found more frequently in goats on dairy operations, while the HR143, NS146, and ND146 genotypes were found more frequently in goats on meat operations. Goats in the east region had a higher percentage of goats with RH154, RQ211, and QK222 genotypes than goats in the west region. The results of this study showed high genetic variability of PRNP among the U.S. goat population, with differences by location and breed, and may serve as a rationale for development of goat breeding programs at the national level to mitigate the risk of scrapie.
... Ataxia and incoordination of movements as well as lumbar itching (scrapie) are the major symptoms of scrapie [44]. Therefore, studies on alterations in the cerebellum have been carried out with special interest. ...
... Both classical scrapie and prion-like diseases induce neurodegeneration, with abundant neuropathological manifestations observable at the level of light and electron microscopy. Four important groups of alterations can be considered: (1) neuronal changes, (2) "spongiform" vesicular (or vacuolar) formations, (3) neuroglial alterations and (4) aberrant prion protein (PrP) deposits/deposition [10][11][12][44][45][46][47] (Figure 1). Therefore, here, we highlight various factors which are extremely relevant. ...
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In this review, the most important neuropathological changes found in the cerebella of sheep affected by classical natural scrapie are discussed. This disease is the oldest known of a group of unconventional “infections” caused by toxic prions of different origins. Scrapie is currently considered a “transmissible spongiform encephalopathy” (due to its neuropathological characteristics and its transmission), which is the paradigm of prion pathologies as well as many encephalopathies (prion-like) that present aberrant deposits of insoluble protein with neurotoxic effects due to errors in their catabolization (“misfolding protein diseases”). The study of this disease is, therefore, of great relevance. Our work data from the authors’ previous publications as well as other research in the field. The four most important types of neuropathological changes are neuron abnormalities and loss, neurogliosis, tissue vacuolization (spongiosis) and pathological or abnormal prion protein (PrP) deposits/deposition. These findings were analyzed and compared to other neuropathologies. Various aspects related to the presentation and progression of the disease, the involution of different neuronal types, the neuroglial responses and the appearance of abnormal PrP deposits are discussed. The most important points of controversy in scrapie neuropathology are presented.
... 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][2][3]. This is supported by the presence of AS in countries that are free of classical scrapie [4,5]. ...
... Small amounts of punctate and granular staining were also seen in the cerebral cortex, basal nuclei, thalamus, and midbrain. In classical scrapie, PrP Sc 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, PrP Sc was observed in the spinal trigeminal tract (Fig 1B), and there was a lack of staining for PrP Sc in the DMNV (Fig 1C). ...
Article
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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.
... Prion diseases are the only zoonotic dementia with such a large host ranges, including scrapie in sheep and goats, bovine spongiform encephalopathy (BSE) in cattle, chronic wasting disease (CWD) in elk and deer, and Creutzfeldt-Jakob disease (CJD), fatal familial insomnia (FFI), and Gerstmann-Sträussler-Scheinker syndrome (GSS) in humans. In CJD, there are three representative types, including sporadic CJD, familial CJD, and iatrogenic CJD [3][4][5][6][7][8][9][10][11][12]. ...
... It has been approximately three centuries since scrapie in sheep was first discovered in 1732 [5]. However, the exact roles on certain factors influencing the causative agent of the disease have not been fully explained thus far. ...
Article
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Prion diseases are caused by misfolded prion protein (PrPSc) and are accompanied by spongiform vacuolation of brain lesions. Approximately three centuries have passed since prion diseases were first discovered around the world; however, the exact role of certain factors affecting the causative agent of prion diseases is still debatable. In recent studies, somatic mutations were assumed to be cause of several diseases. Thus, we postulated that genetically unstable cancer tissue may cause somatic mutations in the prion protein gene (PRNP), which could trigger the onset of prion diseases. To identify somatic mutations in the PRNP gene in cancer tissues, we analyzed somatic mutations in the PRNP gene in cancer patients using the Cancer Genome Atlas (TCGA) database. In addition, to evaluate whether the somatic mutations in the PRNP gene in cancer patients had a damaging effect, we performed in silico analysis using PolyPhen-2, PANTHER, PROVEAN, and AMYCO. We identified a total of 48 somatic mutations in the PRNP gene, including 8 somatic mutations that are known pathogenic mutations of prion diseases. We identified significantly different distributions among the types of cancer, the mutation counts, and the ages of diagnosis between the total cancer patient population and cancer patients carrying somatic mutations in the PRNP gene. Strikingly, although invasive breast carcinoma and glioblastoma accounted for a high percentage of the total cancer patient population (9.9% and 5.4%, respectively), somatic mutations in the PRNP gene have not been identified in these two cancer types. We suggested the possibility that somatic mutations of the PRNP gene in glioblastoma can be masked by a diagnosis of prion disease. In addition, we found four aggregation-prone somatic mutations, these being L125F, E146Q, R151C, and K204N. To the best of our knowledge, this is the first specific analysis of the somatic mutations in the PRNP gene in cancer patients.
... A central assumption for determining RSS associated with genotypes is that scrapie strains do not affect the RSS associated with each genotype. However, different strains of scrapie exist and research results suggest their effect may be genotype dependent [32,37,38]. Nonetheless, results from different countries largely agree on the genotype ranking for RSS [2,4,9]. ...
Article
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Background Scrapie is an infectious prion disease in sheep. Selective breeding for resistant genotypes of the prion protein gene ( PRNP ) is an effective way to prevent scrapie outbreaks. Genotyping all selection candidates in a population is expensive but existing pedigree records can help infer the probabilities of genotypes in relatives of genotyped animals. Results We used linear models to predict allele content for the various PRNP alleles found in Icelandic sheep and compiled the available estimates of relative scrapie susceptibility (RSS) associated with PRNP genotypes from the literature. Using the predicted allele content and the genotypic RSS we calculated estimated breeding values (EBV) for RSS. We tested the predictions on simulated data under different scenarios that varied in the proportion of genotyped sheep, genotyping strategy, pedigree recording accuracy, genotyping error rates and assumed heritability of allele content. Prediction of allele content for rare alleles was less successful than for alleles with moderate frequencies. The accuracy of allele content and RSS EBV predictions was not affected by the assumed heritability, but the dispersion of prediction was affected. In a scenario where 40% of rams were genotyped and no errors in genotyping or recorded pedigree, the accuracy of RSS EBV for ungenotyped selection candidates was 0.49. If only 20% of rams were genotyped, or rams and ewes were genotyped randomly, or there were 10% pedigree errors, or there were 2% genotyping errors, the accuracy decreased by 0.07, 0.08, 0.03 and 0.04, respectively. With empirical data, the accuracy of RSS EBV for ungenotyped sheep was 0.46–0.65. Conclusions A linear model for predicting allele content for the PRNP gene, combined with estimates of relative susceptibility associated with PRNP genotypes, can provide RSS EBV for scrapie resistance for ungenotyped selection candidates with accuracy up to 0.65. These RSS EBV can complement selection strategies based on PRNP genotypes, especially in populations where resistant genotypes are rare.
... Scrapie exists in two forms, the classical form is a contagious disease under natural conditions, while atypical scrapie/Nor98, described for the first time in Norway in 1998, is believed to be a sporadic disease and there is currently no evidence for it being infectious [4]. ...
Article
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Classical scrapie is a contagious prion disease of sheep and goats. It is endemic in many countries in Europe, North America, and Asia. In Africa, imported scrapie cases have been described in South Africa and Kenia in the past. More recently, several cases have been reported from different regions of Libya, based on clinical signs and histological lesions. Here, we report the results of thorough investigations carried out on a suspect case of scrapie in a 6-year-old Barbarine sheep, born, and bred in Tunisia, showing behavioral changes, weight loss, itching, skin lesions, wool loss, and motor incoordination. Histopathology and immunohistochemistry revealed spongiform change in several brain areas with associated pathological prion protein deposition. Western blotting confirmed the diagnosis and showed a classical scrapie-like molecular pattern of PrPres, different from atypical scrapie and bovine spongiform encephalopathy (BSE) in small ruminants. Sequence analysis of the prion protein gene showed that the animal carried the ARQ/ARQ genotype, one of the most susceptible to classical scrapie. The inoculation of sheep brain homogenate in a susceptible rodent model proved the experimental transmissibility of the disease. These results demonstrate the circulation of classical scrapie in Tunisia and confirm its presence in North Africa, indicating the need to improve epidemiological surveillance and diagnostic capacity for prion diseases in the region.
... Many countries have genotype-based eradication programs that emphasize using rams that express arginine at codon 171 in the prion protein, which is associated with resistance to the classical scrapie agent. The US scrapie eradication program has decreased the number of sheep that are scrapie positive at slaughter by 90% [26]. It should be noted that US Meat Animal Research Center is using knowledge of these TMEM154 and PRNP disease resistance genetic variants to develop a Composite IV line of sheep that includes favorable variants for improved OPP and scrapie disease resistance. ...
Article
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Adoption of electronic identification ear tags (EID) and DNA testing by commercial range sheep producers in the Western United States has been low, despite the availability of these technologies for over a decade. Jointly, these technologies offer an approach to provide individual animal performance data to improve flock health, genetic and reproductive management. This project involved a collaboration with five California sheep producers representing a broad geographic range, varying levels of pre-project EID adoption, and diverse operational practices. Tissue samples were collected from, and ear EIDs were placed in, a total of 2,936 rams and their potential lambs. We partnered with a commercial packing company, Superior Farms, to genotype the animals. Superior Farms used a targeted genotyping panel to assign parentage, and link individual animal identification (ID) to camera-graded carcass measurements. This enabled the collection of individual progeny carcass data and provided insight into sire performance, providing for the within-flock identification of prolific sires that were producing lambs with significantly more saleable meat as compared to their flock mates. Overall, almost 91% of lambs were successfully matched to their sire, and prolificacy ranging from 0–135 lambs per ram. There was as much as an $80 difference in the average edible product from camera-graded carcasses derived from lamb groups sired by different rams. A partial budget analysis modeling investment in an EID system coupled with an autodrafter and scale to collect individual weights and improve labor efficiency during processing, and a sheep flip chute to improve worker safety during foot trimmings, yielded a greater than 7:1 return on investment over a five-year time frame. Ideally, the data collection enabled by EIDs and DNA testing would feed into data-driven genetic evaluation programs to enable selection for more productive and profitable animals, and allow the US sheep industry to accelerate the rate of genetic improvement.
... Prion, the causative agent of scrapie, is a protein virus that causes mutations in the central nervous system of sheep, resulting in neuronal death and spongy cavities in brain tissue and the central nervous system. Because no effective treatment is available, once an epidemic of scrapie occurs, the entire flock must be completely culled, which will have a huge impact on the economic benefits of the sheep breeding industry [3,4]. Scrapie has plagued humans for a long time, and according to the relevant literature, scrapie frequently occurred in the European continent before the 18th century [5], with definite records of scrapie starting in Spain in 1732 [6]. ...
Article
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Classical scrapie is a transmissible spongiform encephalopathy that attacks the central nervous system of sheep and goats. Since its discovery in the 18th century, the disease has caused enormous economic losses and public health impacts in continental Europe. In the late 20th century, classical scrapie began to spread to places, such as Asia and the Americas, becoming a disease of global concern. In this study, based on prion occurrence records and high-resolution environmental layers, a risk assessment of classical scrapie in China was performed using a maximum entropy model. The model achieved an area under the curve value of 0.906 (95% confidence interval, 0.0883–0.0929). Sheep distribution density, road density, goat distribution density, minimum temperature of the coldest month, port density, and precipitation of the driest quarter were identified as important factors affecting the occurrence of classical scrapie. The risk map showed that potential high-risk areas in China were mainly located in Northeast China, North China, and Northwest China. This study can provide a valuable reference for the prevention of classical scrapie in China. According to the environmental variables and risk areas of classical scrapie, implementing monitoring and early warning measures in these areas is recommended to reduce the possibility of classical scrapie occurrence and transmission.
... Не існує методів боротьби з атиповою формою скрепі, яка реєструється спорадично й на низькому рівні, і, ймовірно, не розповсюджується легко між тваринами в польових умовах (Greenlee, 2019). ...
Article
У статті наведено аналіз сучасної епізоотичної ситуації щодо небезпечного пріонного захворювання овець, кіз і муфлонів – скрепі. Проаналізовані ветеринарні аспекти охорони здоров’я з урахуванням зоонозного потенціалу цього збудника. Наведено sc характеристику збудника хвороби – інфекційного пріону (PrP ), який є одним із етіологічних агентів групи трансмісивних губчастоподібних енцефалопатій, власне нейродегенеративних захворювань, спричинених пріонами, інфекційними білками, які реплікуються шляхом перетворення нормального клітинного білка в копії зміненого пріона. Наведені сучасні відомості про шляхи розповсюдження збудника та сприйнятливість до скрепі певних порід овець. Особливу увагу в статті присвячено діагностиці захворювання, а саме сучасним методам лабораторних досліджень. Детально описані клінічні ознаки і перебіг класичної та атипової форм скрепі, наведено порівняльну характеристику цих двох форм перебігу. Із урахуванням світового досвіду, висвітнені основні складові, на які потрібно звертати увагу ветеринарним працівникам під час організації заходів боротьби та профілактики цього захворювання. На основі аналітичного дослідження автори роблять висновок про те, що існує нагальна необхідність впровадження активної системи епіднагляду серед овець і кіз з метою виявлення випадків хвороби, що у свою чергу матиме рішуче значення для попередження занесення і розповсюдження хвороби на території України та раннього виявлення спалахів хвороби.
... Transmissible spongiform encephalopathies (TSE) are a disease caused by an infectious protein called a prion. Exposure to a prion in sheep can lead to Scrapie, a neurodegenerative disease [33]. There have been two recorded instances of a European veterinary vaccine leading to the spread of scrapie within vaccinated sheep [34]. ...
Article
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Preventing adventitious agents from contaminating pharmaceutical products has been an important goal of regulatory agencies and industry for decades. Contamination of these products does not only erode consumer trust but also can have potentially serious health consequences. There are a wide variety of adventitious agents that can contaminate many different classifications of products, with each combination requiring different techniques for prevention or detection of adventitious agent contamination. This review seeks to give a brief overview of adventitious agents that have contaminated released pharmaceutical products, explain the different products that are at risk of contamination, then describe the methods commonly used for the prevention and detection of adventitious agent contamination.
... Scrapie is the second most common prion disease that affects sheep and goats that is transferred to humans via contaminated food produce. It is a protein misfolding illness in which the normal prion protein misfolds into a pathogenic form and eventually causes neurodegeneration [27]. While it is known that Scrapie has infected sheep for centuries, cases involving goats are very minimal [28]. ...
... In goats, around 50 polymorphisms of the PRNP gene have been described. However, it has not been determined with the same precision as in sheep if there is a direct correlation between goat genotypes and the susceptibility of contracting scrapie (Greenlee, 2019). For this reason, no breeding programs have been developed in goats, as in the case of sheep. ...
Article
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Animal prion diseases are a group of neurodegenerative, transmissible, and fatal disorders that affect several animal species. The causative agent, prion, is a misfolded isoform of normal cellular prion protein, which is found in cells with higher concentration in the central nervous system. This review explored the sources of infection and different natural transmission routes of animal prion diseases in susceptible populations. Chronic wasting disease in cervids and scrapie in small ruminants are prion diseases capable of maintaining themselves in susceptible populations through horizontal and vertical transmission. The other prion animal diseases can only be transmitted through food contaminated with prions. Bovine spongiform encephalopathy is the only animal prion disease considered zoonotic. However, due to its inability to transmit within a population, it could be controlled. The emergence of atypical cases of scrapie and bovine spongiform encephalopathy, even the recent report of prion disease in camels, demonstrates the importance of understanding the transmission routes of prion diseases to take measures to control them and to assess the risks to human and animal health.
... Unlike CSc, outbreaks of ASc appear to be spontaneous. In addition, ASc tends to disseminate poorly within a flock [1,12]. ...
Article
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Portugal was among the first European countries to report cases of Atypical Scrapie (ASc), the dominant form of Transmissible Spongiform Encephalopathy (TSE) in Portuguese small ruminants. Although the diagnostic phenotypes observed in Portuguese ASc cases seem identical to those described for Nor98, unequivocal identification requires TSE strain-typing using murine bioassays. In this regard, we initiated characterization of ASc isolates from sheep either homozygous for the ARQ genotype or the classical scrapie-resistant ARR genotype. Isolates from such genotypes were transmitted to TgshpXI mice expressing ovine PrPARQ. Mean incubation periods were 414 ± 58 and 483 ± 107 days in mice inoculated with AL141RQ/AF141RQ and AL141RR/AL141RR sheep isolates, respectively. Both isolates produced lesion profiles similar to French ASc Nor98 ‘discordant cases’, where vacuolation was observed in the hippocampus (G6), cerebral cortex at the thalamus (G8) level, cerebellar white matter (W1) and cerebral peduncles (W3). Immunohistochemical PrPSc deposition was observed in the hippocampus, cerebellar cortex, cerebellar white matter and cerebral peduncles in the form of aggregates and fine granules. These findings were consistent with previously reported cases of ASc Nor98 transmitted to transgenic TgshpXI mice, confirming that the ASc strain present in Portuguese sheep corresponds to ASc Nor98.
... Cases of atypical scrapie are distinguished by a 5-band profile on Western blot with a prominent lower band at approximately 11 to 12 kDa, whereas, classical scrapie has the nonglycosylated band at approximately 19 to 21 kDa. In addition, the PrP Sc of atypical scrapie is relatively sensitive to proteinases [30,31]. In the last 15 years, many polymorphisms associated with resistance/susceptibility to scrapie, such as 110T/P, 127G/S, 142M/I, 145G/D, 146N/D/S, 154R/H, 211R/Q, and 222Q/K have been described in different breeds of goats and regions [5,[32][33][34][35][36][37][38][39][40]. ...
Article
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Scrapie is considered an endemic disease in both sheep and goats in Greece. However, contrary to sheep, in goats more than one prion protein (PrP) polymorphism has been recognized as a candidate for resistance breeding against the disease. For an impression which candidates are circulating, (i) brain samples (n = 525) from scrapie-affected (n = 282) and non-affected (n = 243) animals within the national surveillance program, and (ii) individual blood samples (n = 1708) from affected (n = 241) and non-affected (n = 1467) herds, in a large part of mainland Greece and its islands, were collected and assayed. A dedicated Taqman method was used to test for amino acid polymorphisms 110T/P, 146N/S/D, 211R/Q, and 222Q/K. Highly prevalent genotypes were 110TT, 146NN, 211RR, and 222QQ. The frequencies of polymorphisms in blood and negative brain samples for codons 110P, 211Q, and 222K were 4.0%, 3.0%, and 1.9%, respectively, while 146D (0.7%) was present only on Karpathos island. Codon 110P was exclusively found in scrapie-negative brains, and homozygous 110P/P in two scrapie-negative goats. It is concluded that breeding programs in Karpathos could focus on codon 146D, while in other regions carriers of the 110P and 222K allele should be sought. Case-control and challenge studies are now necessary to elucidate the most efficient breeding strategies.
... Scrapie is a prion disease of sheep, endemic in many countries including the United Kingdom, which comprises several distinct strains (Greenlee, 2019). There is polymorphic variation in the sheep PRNP at amino acid positions 136, 154, and 171 that affects susceptibility to scrapie, and the VRQ, ARQ, and ARR genotypes have the greatest effect in this regard (Goldmann, 2008;Gonzalez et al., 2012). ...
Article
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Prion diseases are fatal neurodegenerative disorders that affect humans and animals, and can also be transmitted from animals to humans. A fundamental event in prion disease pathogenesis is the conversion of normal host prion protein (PrPC) to a disease-associated misfolded form (PrPSc). Whether or not an animal prion disease can infect humans cannot be determined a priori. There is a consensus that classical bovine spongiform encephalopathy (C-type BSE) in cattle transmits to humans, and that classical sheep scrapie is of little or no risk to human health. However, the zoonotic potential of more recently identified animal prion diseases, such as atypical scrapie, H-type and L-type BSE and chronic wasting disease (CWD) in cervids, remains an open question. Important components of the zoonotic barrier are (i) physiological differences between humans and the animal in question, (ii) amino acid sequence differences of the animal and human PrPC, and (iii) the animal prion strain, enciphered in the conformation of PrPSc. Historically, the direct inoculation of experimental animals has provided essential information on the transmissibility and compatibility of prion strains. More recently, cell-free molecular conversion assays have been used to examine the molecular compatibility on prion replication and zoonotic potential. One such assay is Protein Misfolding Cyclic Amplification (PMCA), in which a small amount of infected tissue homogenate, containing PrPSc, is added as a seed to an excess of normal tissue homogenate containing PrPC, and prion conversion is accelerated by cycles of incubation and ultrasonication. PMCA has been used to measure the molecular feasibility of prion transmission in a range of scenarios using genotypically homologous and heterologous combinations of PrPSc seed and PrPC substrate. Furthermore, this method can be used to speculate on the molecular profile of PrPSc that might arise from a zoonotic transmission. We discuss the experimental approaches that have been used to model both the intra- and inter-species molecular compatibility of prions, and the factors affecting PrPc to PrPSc conversion and zoonotic potential. We conclude that cell-free prion protein conversion assays, especially PMCA, are useful, rapid and low-cost approaches for elucidating the mechanisms of prion propagation and assessing the risk of animal prions to humans.
... AS is different from CS regarding its clinical presentation, but also in its molecular characteristics, the anatomical distribution of PrP Sc within infected sheep, the genotypes affected and its epidemiology (see Greenlee, 2019, for recent review). Whereas more than 20 different strains of CS have been described, only one has been demonstrated in AS so far (Ac ın et al., 2021). ...
Article
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The European Commission asked EFSA whether the scientific data on the 2-year intensified monitoring in atypical scrapie (AS) outbreaks (2013-2020) provide any evidence on the contagiousness of AS, and whether they added any new knowledge on the epidemiology of AS. An ad hoc data set from intensified monitoring in 22 countries with index case/s of AS in sheep and/or goats (742 flocks from 20 countries, 76 herds from 11 countries) was analysed. No secondary cases were confirmed in goat herds, while 35 secondary cases were confirmed in 28 sheep flocks from eight countries. The results of the calculated design prevalence and of a model simulation indicated that the intensified monitoring had limited ability to detect AS, with no difference between countries with or without secondary cases. A regression model showed an increased, but not statistically significant, prevalence (adjusted by surveillance stream) of secondary cases in infected flocks compared with that of index cases in the non-infected flocks (general population). A simulation model of within-flock transmission, comparing a contagious (i.e. transmissible between animals under natural conditions) with a non-contagious scenario, produced a better fit of the observed data with the non-contagious scenario, in which each sheep in a flock had the same probability of developing AS in the first year of life. Based on the analyses performed, and considering uncertainties and data limitations, it was concluded that there is no new evidence that AS can be transmitted between animals under natural conditions, and it is considered more likely (subjective probability range 50-66%) that AS is a non-contagious, rather than a contagious disease. The analysis of the data of the EU intensified monitoring in atypical scrapie infected flocks/herds confirmed some of the known epidemiological features of AS but identified that major knowledge gaps still remain.
... After intracranial inoculation, VRQ/ARQ and ARQ/ARQ sheep had widespread lymphoid distribution of PrP Sc that was reminiscent of PrP Sc distribution in natural scrapie infection. 6,14 However, ARQ/ARR sheep did not have detectable lymphoid PrP Sc with IHC. The overall absence of lymphoid accumulation in the ARQ/ARR sheep is similar to ARQ/ARR sheep inoculated intracranially with the scrapie agent. ...
Article
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The origin of chronic wasting disease (CWD) in cervids is unclear. One hypothesis suggests that CWD originated from scrapie in sheep. We compared the disease phenotype of sheep-adapted CWD to classical scrapie in sheep. We inoculated sheep intracranially with brain homogenate from first-passage mule deer CWD in sheep (sCWD md ). The attack rate in second-passage sheep was 100% (12 of 12). Sheep had prominent lymphoid accumulations of PrP Sc reminiscent of classical scrapie. The pattern and distribution of PrP Sc in the brains of sheep with CWD md was similar to scrapie strain 13-7 but different from scrapie strain x124. The western blot glycoprofiles of sCWD md were indistinguishable from scrapie strain 13-7; however, independent of sheep genotype, glycoprofiles of sCWD md were different than x124. When sheep genotypes were evaluated individually, there was considerable overlap in the glycoprofiles that precluded significant discrimination between sheep CWD and scrapie strains. Our data suggest that the phenotype of CWD in sheep is indistinguishable from some strains of scrapie in sheep. Given our results, current detection techniques would be unlikely to distinguish CWD in sheep from scrapie in sheep if cross-species transmission occurred naturally. It is unknown if sheep are naturally vulnerable to CWD; however, the susceptibility of sheep after intracranial inoculation and lymphoid accumulation indicates that the species barrier is not absolute.
... Scrapie is a transmissible spongiform encephalopathy (TSE) that occurs in sheep and goats. It is a neurodegenerative disease characterised by abnormal behaviour, lip-smacking, blindfolding, altered gaits and dysmetria (Greenlee, 2019;Kim et al., 2019aKim et al., , 2019bCassmann and Greenlee, 2020). TSE includes chronic wasting disease (CWD) in elk and deer, bovine spongiform encephalopathy (BSE) in cattle, feline spongiform encephalopathy (FSE) in cheetahs, pumas and cats, and Creutzfeldt-Jakob disease (CJD) in humans (Imran and Mahmood, 2011;Jeong and Kim, 2014;Benestad and Telling, 2018;Kim and Jeong, 2018;Houston and Andreoletti, 2019;Won et al., 2020). ...
Article
Prion disease is a fatal neurodegenerative disease with a broad host range in humans and animals. It is caused by proteinase K-resistant prion protein (PrP res ). In previous studies, a heterogeneous infection in Cervidae and Caprinae was reported. Chronic wasting disease (CWD) has been frequently reported as the only prion disease in Korea that occurs in livestock. Thus, there is a possibility of transmission of CWD to Korean native black goats. However, PrP res has not been investigated thus far in Korean native black goats. We found strong linkage disequilibrium between c.126G>A and c.414T>C ( r ² = 1) and between c.718C>T and c.126G>A ( r ² = 0.638). In addition, the haplotype GTGTAAAC (representing codons 42, 102, 127, 138, 143, 146, 218 and 240) showed the highest frequency with 45.1%. Among 41 Korean native black goats, 20 animals (48.78%) were homozygous for the susceptible haplotypes (histidine at codon 143, asparagine at codon 146 and arginine at codon 154). Interestingly, we did not detect PrP res bands in any of the tested animals, including the 20 animals carrying potential scrapie susceptible haplotypes.
... Prion diseases are caused by misfolded prion protein (PrP Sc ) and denote paramount infection ranges and include various types, such as Creutzfeldt-Jakob disease (CJD) in humans, scrapie in sheep and goats, bovine spongiform encephalopathy (BSE) in cattle and chronic wasting disease (CWD) in elk and deer [1][2][3][4][5][6][7][8][9]. Recent studies have reported that the susceptibility of prion disease was affected by polymorphisms of the prion gene family, including prion protein gene (PRNP), prion-like protein gene (PRND) and shadow of prion protein (SPRN) [10][11][12][13][14][15][16][17][18]. ...
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Chronic wasting disease (CWD) is caused by abnormal deleterious prion protein (PrPSc), and transmissible spongiform encephalopathy occurs in the Cervidae family. In recent studies, the susceptibility of prion disease has been affected by polymorphisms of the prion gene family. However, the study of the prion-related protein gene (PRNT) is rare, and the DNA sequence of this gene was not fully reported in all Cervidae families. In the present study, we amplified and first identified PRNT DNA sequences in the Cervidae family, including red deer, elk, sika deer and Korean water deer, using polymerase chain reaction (PCR). We aligned nucleotide sequences of the PRNT gene and the amino acid sequences of prion-related protein (Prt) protein among several species. In addition, we performed phylogenetic analysis to measure the evolutionary relationships of the PRNT gene in the Cervidae family. Furthermore, we performed homology modeling of the Prt protein using SWISS-MODEL and compared the structure of Prt protein between sheep and the Cervidae family using the Swiss-PdbViewer program. We obtained much longer PRNT sequences of red deer compared to the PRNT gene sequence registered in GenBank. Korean water deer denoted more close evolutionary distances with goats and cattle than the Cervidae family. We found 6 Cervidae family-specific amino acids by the alignment of Prt amino acid sequences. There are significantly different distributions of hydrogen bonds and the atomic distance of the N-terminal tail and C-terminal tail between sheep and the Cervidae family. We also detected the mRNA expression of PRNT gene in 3 tissues investigated. To our knowledge, this report is the first genetic study of the PRNT gene in the Cervidae family.
... Pathological phenotypic differences between atypical and classical scrapie have been reviewed. 157 Unlike sheep with classical scrapie, PrP Sc is undetectable by immunoassay in the lymphoid tissues of sheep with atypical scrapie. However, sheep with atypical scrapie were determined to have infectious PrP Sc in lymphoid tissue, muscles, and peripheral ner-vous tissue on the basis of positive bioassay results in transgenic mice that overexpress the sheep prion protein. ...
Article
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In sheep, scrapie is a fatal neurologic disease that is caused by a misfolded protein called a prion (designated PrPSc). The normal cellular prion protein (PrPC) is encoded by an endogenous gene, PRNP, that is present in high concentrations within the CNS. Although a broad range of functions has been described for PrPC, its entire range of functions has yet to be fully elucidated. Accumulation of PrPSc results in neurodegeneration. The PRNP gene has several naturally occurring polymorphisms, and there is a strong correlation between scrapie susceptibility and PRNP genotype. The cornerstone of scrapie eradication programs is the selection of scrapie-resistant genotypes to eliminate classical scrapie. Transmission of classical scrapie in sheep occurs during the prenatal and periparturient periods when lambs are highly susceptible. Initially, the scrapie agent is disseminated throughout the lymphoid system and into the CNS. Shedding of the scrapie agent occurs before the onset of clinical signs. In contrast to classical scrapie, atypical scrapie is believed to be a spontaneous disease that occurs in isolated instances in older animals within a flock. The agent that causes atypical scrapie is not considered to be naturally transmissible. Transmission of the scrapie agent to species other than sheep, including deer, has been experimentally demonstrated as has the transmission of nonscrapie prion agents to sheep. The purpose of this review is to outline the current methods for diagnosing scrapie in sheep and the techniques used for studying the pathogenesis and host range of the scrapie agent. Also discussed is the US scrapie eradication program including recent updates.
... Prion diseases are a group of fatal and infectious neurodegenerative disorders that include Creutzfeldt-Jacob disease (CJD) in humans, bovine spongiform encephalopathy (BSE) in cattle, scrapie in sheep and goat, and chronic wasting disease (CWD) in cervids. Disease manifestation in human prion disorders can be sporadic, familial, or acquired by infection (1)(2)(3)(4)(5), whereas in animals acquired prion diseases prevail (6)(7)(8)(9). Pathological hallmarks of prion diseases are vacuolation caused by neuronal death, PrP Sc plaque deposition, astrogliosis and spongiform degeneration (10). Prion diseases are caused by PrP Sc , the pathological isoform of the normal cellular prion protein (PrP C ), which is highly expressed in neurons, astrocytes, and oligodendrocytes (11). ...
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Prion diseases are fatal infectious neurodegenerative disorders in human and animals caused by misfolding of the cellular prion protein (PrP C ) into the pathological isoform PrP Sc . Elucidating the molecular and cellular mechanisms underlying prion propagation may help to develop disease interventions. Cell culture systems for prion propagation have greatly advanced molecular insights into prion biology, but translation of in vitro to in vivo findings is often disappointing. A wider range of cell culture systems might help overcome these shortcomings. Here, we describe an immortalized mouse neuronal astrocyte cell line (C8D1A) that can be infected with murine prions. Both PrP C protein and mRNA levels in astrocytes were comparable to those in neuronal and nonneuronal cell lines permitting persistent prion infection. We challenged astrocytes with three mouse-adapted prion strains (22L, RML, and ME7) and cultured them for six passages. Immunoblotting results revealed that the astrocytes propagated 22L prions well over all six passages, whereas ME7 prions did not replicate, and RML prions only very weakly after five passages. Immunofluorescence analysis indicated similar results for PrP Sc . Interestingly, when we used prion conversion activity as a read-out in real-time quaking-induced conversion (RT-QuIC) assays with RML-infected cell lysates, we observed a strong signal over all six passages, comparable to that for 22L-infected cells. These data indicate that the C8D1A cell line is permissive to prion infection. Moreover, the propagated prions differed in conversion and proteinase K–resistance levels in these astrocytes. We propose that the C8D1A cell line could be used to decipher prion strain biology.
... More than 50 polymorphisms in the goat prion protein-coding gene were reported worldwide 25 . A recent study on Ethiopian goat population reported six variants in Western Highland, Central Highland, and Long Eared Somali breeds of Ethiopia Table 5 Content courtesy of Springer Nature, terms of use apply. ...
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Scrapie is a lethal neurodegenerative disease of sheep and goats caused by the misfolding of the prion protein. Variants such as M142, D145, S146, H154, Q211, and K222 were experimentally found to increase resistance or extend scrapie incubation period in goats. We aimed to identify polymorphisms in the Afar and Arsi-Bale goat breeds of Ethiopia and computationally assess the effect of variants on prion protein stability. In the present study, four non-synonymous novel polymorphisms G67S, W68R, G69D, and R159H in the first octapeptide repeat and the highly conserved C-terminus globular domain of goat PrP were detected. The resistant genotype, S146, was detected in >50% of the present population. The current study population showed a genetic diversity in Ethiopian goat breeds. In the insilico analysis, the R68 variant was predicted to increase stability while S67, D69, and H159 decrease the stability of prion protein. The new variants in the octapeptide repeat motif were predicted to decrease amyloidogenicity but H159 increased the hotspot sequence amyloidogenic propensity. These novel variants could be the source of conformational flexibility that may trigger the gain or loss of function by prion protein. Further experimental study is required to depict the actual effects of variants on prion protein stability.
Article
Amyloidosis is a group of diseases in which proteins become amyloid, an insoluble fibrillar aggregate, resulting in organ dysfunction. Amyloid deposition has been reported in various animal species. To diagnose and understand the pathogenesis of amyloidosis, it is important to identify the amyloid precursor protein involved in each disease. Although 42 amyloid precursor proteins have been reported in humans, little is known about amyloidosis in animals, except for a few well-described amyloid proteins, including amyloid A (AA), amyloid light chain (AL), amyloid β (Aβ), and islet amyloid polypeptide-derived amyloid. Recently, several types of novel amyloidosis have been identified in animals using immunohistochemistry and mass spectrometry–based proteomic analysis. Certain species are predisposed to specific types of amyloidosis, suggesting a genetic background for its pathogenesis. Age-related amyloidosis has also emerged due to the increased longevity of captive animals. In addition, experimental studies have shown that some amyloids may be transmissible. Accurate diagnosis and understanding of animal amyloidosis are necessary for appropriate therapeutic intervention and comparative pathological studies. This review provides an updated classification of animal amyloidosis, including associated protein misfolding disorders of the central nervous system, and the current understanding of their pathogenesis. Pathologic features are presented together with state-of-the-art diagnostic methods that can be applied for routine diagnosis and identification of novel amyloid proteins in animals.
Article
Three‐dimensional morphometric data better show the structural and functional characteristics of the brain. The objective of this study was to estimate the volume of the cerebral structures of the sheep using design‐based stereology. The brains of five sheep were used, fixed in formalin 10% and embedded in agar 6%. An average of 10–12 slab was obtained from each brain. All slabs were stained using Mulligan's method and photographs were recorded. The volume of the brain and its structures were estimated using the Cavalieri's estimator and the point counting system. The total volume was 70604.8 ± 132.45 mm ³ . The volume fractions of the grey and white matters were calculated as 42.55 ± 0.21% and 24.23 ± 0.51% of the whole brain, respectively. The fractional volume of the caudate nucleus and claustrum were estimated at 2.39 ± 0.08% and at 1.008 ± 0.057% of total brain volume. The volumes of corpus callosum, internal capsule and external capsule were 1.24 ± 0.053%, 3.63 ± 0.22% and 0.698 ± 0.049% of total cerebral volume, respectively. These data could help improve the veterinary comparative neuroanatomy knowledge and development of experimental studies in the field.
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Background Prion diseases, also known as transmissible spongiform encephalopathies (TSEs) remain one of the deleterious disorders, which have affected several animal species. Polymorphism of the prion protein (PRNP) gene majorly determines the susceptibility of animals to TSEs. However, only limited studies have examined the variation in PRNP gene in different Nigerian livestock species. Thus, this study aimed to identify the polymorphism of PRNP gene in Nigerian livestock species (including camel, dog, horse, goat, and sheep). We sequenced the open reading frame (ORF) of 65 camels, 31 village dogs and 12 horses from Nigeria and compared with PRNP sequences of 886 individuals retrieved from public databases. Results All the 994 individuals were assigned into 162 haplotypes. The sheep had the highest number of haplotypes (n = 54), and the camel had the lowest (n = 7). Phylogenetic tree further confirmed clustering of Nigerian individuals into their various species. We detected five non-synonymous SNPs of PRNP comprising of G9A, G10A, C11G, G12C, and T669C shared by all Nigerian livestock species and were in Hardy-Weinberg Equilibrium (HWE). The amino acid changes in these five non-synonymous SNP were all “benign” via Polyphen-2 program. Three SNPs G34C, T699C, and C738G occurred only in Nigerian dogs while C16G, G502A, G503A, and C681A in Nigerian horse. In addition, C50T was detected only in goats and sheep. Conclusion Our study serves as the first to simultaneously investigate the polymorphism of PRNP gene in Nigerian livestock species and provides relevant information that could be adopted in programs targeted at breeding for prion diseases resistance.
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Scrapie is a contagious disease of sheep and goats caused by prions (PrPSc). This study described an outbreak of Scrapie in sheep in the state of Santa Catarina, Brazil. An 1-year and 3-month-old sheep developed clinical signs characterized by motor incoordination of the pelvic limbs, pruritus and alopecia for three days. The 38 sheep from the flock that were over 1 year of age underwent biopsies of the third eyelid and rectal mucosa, in addition to anti-PrPsc immunohistochemistry (IHC). Blood containing EDTA was collected for PRNP gene genotyping from these sheep. Of the 38, 16 (42.10%) had immunostaining againstPrPSc. IHC-positive animals were euthanized and necropsied, as well as lambs from positive mothers. Different organs of the 19 necropsied animals were collected in 10% buffered formalin for histopathological examination and anti-PrPSc IHC of the obex. The histopathology of the obex of the female with neurological signs presented discrete multifocal vacuolization of the cytoplasm of neurons and neuropil. The anti-PrPSc IHC showed that two out of the 19 obex samples had cytoplasmic immunostaining in neurons. The genotypes reported were ARQ/ARQ in 47.36%, ARR/ARQ in 36.84%, ARQ/VRQ in 10.52% and ARQ/VRR in 5.28%. The genotyping helps to identify susceptible animals and select animals more resistant to the development of Scrapie. The anti-PrPSc IHC from lymphoid biopsies, and genotyping demonstrated the high number of positive sheep classified in susceptible group.
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У науковій монографії викладено сучасні погляди на пріони та пріонні хвороби тварин і людини. Автори, ґрунтуючись на наукових працях і сучасних розробках щодо діагностики, розповсюдження і профілактики пріонних захворювань розкривають значення цих хвороб в епізоотології й епідеміології, наводять загальні закономірності й тенденції їх поширення, розкривають сучасні відомості про пріони, розкривають питання зоонозного потенціалу пріонних захворювань. Наведено характеристики й особливості перебігу пріонних хвороб тварин і людини, описані епізоотологічні й епідеміологічні їх особливості, викладено сучасні методи діагностики (у тому числі методи білкової ампліфікації) й заходи захисту від цих хвороб. Наукова монографія розрахована на спеціалістів районних і обласних управлінь, лікарень ветеринарної медицини, слухачів інститутів і факультетів післядипломного навчання, викладачів та студентів факультетів ветеринарної медицини вищих навчальних аграрних закладів і загал практичних фахівців ветеринарної медицини. The scientific monograph presents current views on prions and animal and human prion diseases. Authors based on scientific works and modern developments in diagnosis, distribution and prevention of prion diseases reveal the importance of these diseases in epizootology and epidemiology, state the general patterns and trends of their distribution, reveal modern information about prions, reveal questions zoonotic potential of prion diseases. The scientific monograph is intended for district and regional specialists departments, hospitals of veterinary medicine, students of institutes and faculties of postgraduate studies, teachers and students of faculties of veterinary medicine of higher educational agrarian institutions and general practitioners of veterinary medicine.
Chapter
This chapter presents most of the neurologic diseases of goats, with some exceptions. It discusses the neurologic aspects of caprine arthritis encephalitis in conjunction with maedi‐visna, a closely related retroviral disease. The chapter presents the clinical signs associated with the lesions in different regions of the nervous system. Scrapie is an infectious, contagious, degenerative neurologic disease that occurs naturally only in sheep and goats. Border disease, also known as “hairy shaker” disease in lambs, is an infectious, contagious viral disease of sheep and goats. Louping‐ill, also known as ovine encephalomyelitis, is an acute, non‐contagious encephalomyelitis caused by the louping‐ill virus, an enveloped, positive‐sense, single‐stranded ribonucleic acid virus in the genus Flavivirus . Spastic Paresis is well known in cattle and is considered to be inherited in that species.
Chapter
The infectious disorders are grouped as bacterial, viral, protozoal, fungal, amoebic, verminous and chlamydial. These are followed by the Inflammatory and Immune‐related disorders the granulomatous CNS diseases, idiopathic eosinophilic encephalomyelitides and sclerosing panencephalitis then by the presumed junctional disorder myasthenia gravis. Biotoxicoses are covered in Chapter 34. As in other chapters in Part III the diseases are ordered to follow the anatomic groupings: diffuse brain, forebrain, brainstem, cerebellum, spinal cord, peripheral nerves, neuromuscular junction, muscle.
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Transmisivne spongiformne encefalopatije su grupa rijetkih, progresivnih neurodegenerativnih bolesti od kojih obolijevaju ljudi i veći broj životinjskih vrsta, a prouzroči ih prion. Infekcija je dugi vremenski period asimptomatska, a bolest je progresivna i neminovno letalnog ishoda. U skupinu transmisivnih spongiformnih encefalopatija spadaju: klasična i atipična goveđa spongiformna encefalopatija, klasični i atipični grebež u ovaca i koza, spongiformna encefalopatija jelena, transmisivna encefalopatija američke vidrice, mačja spongiformna encefalopatija, Creutzfeldt- Jakobova bolest, varijanta Creutzfeldt- Jakobove bolesti, Gerstmann-Sträussler- Scheinker sindrom, familijarna fatalna insomnija i kuru. Za postojanje ove bolesti na ovcama zna se već gotovo tri stoljeća. Predmetne bolesti su se izučavale tijekom 20. stoljeća, ali su se tek pojavom epizootije goveđe spongiformne encefalopatije 80-ih godina prošlog stoljeća u Ujedinjenom Kraljevstvu (UK) te otkrivanjem novih dijagnostičkih metoda, istraživanja intenzivirala. Otkriven je i uzročnik bolesti - prion (proteinska zarazna čestica). Prion je normalni stanični protein pronađen prije svega na površini neurona, ali i drugih stanica u organizmu, koji se zbog još uvijek neobjašnjivih razloga u biokemijskim procesima sinteze proteina promijeni i postaje uzročnik encefalopatije. U ovome članku prikazani su rezultati praćenja goveđe spongiformne encefalopatije i grebeža ovaca u Republici Hrvatskoj od 2001. do kraja 2021. godine. Goveđa spongiformna encefalopatija se sistematski nadzire od 2001., dok se grebež ovaca nadzire od 2002. godine. Do kraja 2021. godine ukupno je pretraženo 454.822 uzoraka moždanog tkiva goveda, a tijekom navedenog perioda nije utvrđen niti jedan pozitivan uzorak. Na grebež ovaca pretraženo je ukupno 25.332 uzoraka moždanog tkiva koza i ovaca. Laboratorijskim pretraživanjem nije utvrđen niti jedan pozitivan uzorak na klasični grebež ovaca, ali su potvrđena četiri slučaja atipičnog grebeža u ovaca. Na temelju svih provedenih aktivnosti i nadziranja goveđe spongiformne encefalopatije u Republici Hrvatskoj je od strane Svjetske organizacije za zdravlje životinja 2014. godine dodijeljen status zemlje sa zanemarivim rizikom u odnosu na goveđu spongiformnu encefalopatiju, što je ujedno i najviši zdravstveni status u odnosu na ovu bolest.
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Livestock is heavily affected by trypanosomosis in Africa. Through strong selective pressure, several African indigenous breeds of cattle and small ruminants have acquired varying degrees of tolerance against this disease. In this study, we combined LFMM and PCAdapt for analyzing two datasets of goats from West-Central Africa and East Africa, respectively, both comprising breeds with different assumed levels of trypanotolerance. The objectives were (i) to identify molecular signatures of selection related to trypanotolerance; and (ii) to guide an optimal sampling for subsequent studies. From 33 identified signatures, 18 had been detected previously in the literature as being mainly associated with climatic adaptations. The most plausible signatures of trypanotolerance indicate the genes DIS3L2, COPS7B, PD5A, UBE2K, and UBR1. The last gene is of particular interest since previous literature has already identified E3-ubiquitin ligases as playing a decisive role in the immune response. For following-up on these findings, the West-Central African area appears particularly relevant because of (i) a clear parasitic load gradient related to a humidity gradient, and (ii) still restricted admixture levels between goat breeds. This study illustrates the importance of protecting local breeds, which have retained unique allelic combinations conferring their remarkable adaptations.
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The formation of amyloid fibrils is critical for neurodegenerative diseases. Some physiochemical conditions can promote the conversion of proteins from soluble globular shapes into insoluble well‐organized amyloid fibrils. The aim of this study was to investigate the effect of temperatures on amyloid fibrils formation in vitro using the protein model of hen egg‐white lysozyme (HEWL). The HEWL fibrils were prepared at temperatures of 37, 45, 50 and 57°C in glycine solution of pH 2.2. Under transmission electron microscopy, we found the well‐organized HEWL amyloid fibrils at temperatures of 45, 50 and 57°C after 10 days of incubation. Thioflavin T and Congo red florescence assays confirmed that the formation and growth of HEWL fibrils displayed a temperature‐dependent increase, and 57°C produced the most amounts. Meanwhile, the surface hydrophobicity of aggregates was greatly increased by ANS binding assay, and β‐sheet contents by circular dichroism analysis were increased by 17.8%, 22.0% and 34.9%, respectively. Furthermore, the HEWL fibrils formed at 57°C caused significant cytotoxicity in SH‐SY5Y cells after 48 hr exposure, and the cell viability determined by MTT assay was decreased, with 81.35 ± 0.29% for 1 μM, 61.45 ± 2.62% for 2 μM, and 11.58 ± 0.39% (p < .01) for 3 μM. Nuclear staining results also confirmed the apoptosis features. These results suggest that the elevated temperatures could accelerate protein unfolding of the native structure and formation of toxic amyloid fibrils, which can improve understanding the mechanisms of the unfolding and misfolding process of prion protein.
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Bighorn sheep (Ovis canadensis) are predicted to have a degree of susceptibility to the transmissible spongiform encephalopathies (TSE) chronic wasting disease and scrapie. We opportunistically screened 127 captive bighorn sheep and 152 free-ranging bighorn sheep in Colorado, US for the presence of TSE over a period of 35 yr. None of the animals demonstrated clinical signs, gross pathology, histopathology, or immunohistochemical staining patterns suggestive of TSE.
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Bovine spongiform encephalopathy (BSE) is a kind of prion disease caused by proteinase K‐resistant prion protein (PrPSc) in cattle. Although BSE has been reported worldwide, BSE‐infected cases have never been reported in Korea. In a previous study, we identified BSE‐related somatic mutation E211K in 3 Korean Holstein cattle. In Korea, the BSE surveillance system has been established. However, several genetic factors have not been controlled simultaneously thus far. In the present study, we performed enhanced surveillance of prion disease‐related factors in Korean cattle, including Holstein cattle and Hanwoo (Korean native cattle), which is widely raised for meat. We investigated the germline mutation E211K at codon 211 of the PRNP gene and analyzed genotype, allele and haplotype frequencies of the 23‐bp and 12‐bp insertion/deletion polymorphisms of the PRNP gene using direct DNA sequencing. In addition, we investigated linkage disequilibrium (LD) and compared haplotype distributions of polymorphisms among cattle breeds. Furthermore, we carried out BSE diagnosis in the medulla oblongata (MO) of Korean cattle including 3 Korean Holstein cattle carrying somatic mutation E211K using western blotting analysis. We did not find the E211K mutation in the PRNP gene in any of the Korean cattle and found significantly different genotype, allele and haplotype distributions of the 23‐bp and 12‐bp insertion/deletion polymorphisms of the PRNP gene in male Holstein compared to male Hanwoo, female Hanwoo and total Hanwoo. In addition, only male Holstein showed weak LD between 23‐bp and 12‐bp insertion/deletion polymorphisms. Furthermore, the PrPSc bands were not detected in all Korean cattle tested. To the best of our knowledge, the enhanced surveillance system of BSE was conducted for the first time in Korean cattle.
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Prions cause transmissible and inevitably fatal neurological diseases in agriculturally important animals, including bovine spongiform encephalopathy in domestic cattle, scrapie in sheep and goats, and chronic wasting disease in cervids. Because animals are largely asymptomatic throughout the course of the disease, early detection of prion disease is important. Hamsters were peripherally (ip) inoculated with hamster-adapted (Sc237) prions. By week 13 of a 14-week disease course, clinical signs appeared. A multiple-reaction-monitoring-based method was used to quantitate the amount of proteinase-K-digested prions (PrP 27-30) and the extent of methionine 213 oxidation present in the brains of infected hamsters. Detectable amounts of PrP 27-30 were present in all animals after 4 weeks. The extent of methionine 213 oxidation decreased over time. When we compared our quantitation results to those from other researchers using bioassay, we observed that consistent detection of PrP 27-30 by mass spectrometry occurs at a time when prions are reliably detected by bioassay.
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The presence of lysine (K) at codon 222 has been associated with resistance to classical scrapie in goats, but few scrapie cases have been identified in 222Q/K animals. To investigate the contribution of the 222K variant to PrPres formation in natural and experimental Q/K scrapie cases, we applied an immunoblotting method based on the use of two different monoclonal antibodies, F99/97.6.1 and SAF84, chosen for their different affinities to 222K and 222Q PrP variants. Our finding that PrPres seems to be formed nearly totally by the 222Q variant provides evidence that the 222K PrP variant confers resistance to conversion to PrPres formation and reinforces the view that this mutation has a protective role against classical scrapie in goats.
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Background. A study to investigate transmission of classical scrapie via goat milk was carried out in sheep: firstly, lambs were challenged orally with goat scrapie brain homogenate to confirm transmission of scrapie from goats to sheep. In the second study phase, milk from scrapie-infected goats was fed to lambs. Lambs were selected according to their prion protein gene (PRNP) genotype, which was either VRQ/VRQ or ARQ/ARQ, with or without additional polymorphisms at codon 141 (FF141, LF141 or LL141) of the ovine PRNP. This report describes the clinical, pathological and molecular phenotype of goat scrapie in those sheep that progressed to clinical end-stage. Results. Ten sheep (six VRQ/VRQ and four ARQ/ARQ, of which three FF141 and one LL141) challenged with one of two scrapie brain homogenates, and six pairs of sheep (ARQ, of which five LL141 and seven LF141) fed milk from six different goats, developed clinical disease, which was characterised by a pruritic (all VRQ/VRQ and LL141 sheep) or a non-pruritic form (all LF141 and FF141 sheep). Immunohistochemical (IHC) examination revealed that the pattern of intra- and extracellular accumulation of disease-associated prion protein in the brain was also dependent on PRNP polymorphisms at codon 141, which was similar in VRQ and LL141 sheep but different from LF141 and FF141 sheep. The influence of codon 141 was also seen in discriminatory Western blot (WB), with LF141 and FF141 sheep showing a bovine spongiform encephalopathy-like profile (diminished reactivity with P4 antibody) on brain tissue. However, discriminatory WB in lymphoid tissues, and IHC pattern and profile both in lymphoid and brain tissue was consistent with classical scrapie in all sheep. Conclusions. This study provided further evidence that the clinical presentation and the pathological and molecular phenotypes of scrapie in sheep are influenced by PRNP polymorphisms, particularly at codon 141. Differences in the truncation of disease-associated prion protein between LL141 sheep and those carrying the F141 allele may be responsible for these observations.
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Scrapie and bovine spongiform encephalopathy (BSE) are transmissible spongiform encephalopathies (TSE's) affecting sheep and goats. Susceptibility of goats to scrapie is influenced by polymorphisms of the prion protein gene (PRNP) of the host. Five polymorphisms are associated with reduced susceptibility to TSE's. In the study presented here caprine samples from a scrapie eradication program on Cyprus were genotyped and further characterized using BioRad TeSeE rapid test, histological, immunohistochemical and biochemical methods. In total 42 goats from 20 flocks were necropsied from which 25 goats showed a positive result in the rapid test, a spongiform encephalopathy and an accumulation of pathological prion protein (PrPSc) in the obex. PrPSc deposits were demonstrated in the placenta, peripheral nervous and lymphoreticular system. Two animals showed PrPSc-accumulations in peripheral tissues only. By discriminatory immunoblots a scrapie infection could be confirmed for all cases. Nevertheless, slight deviations in the glycosylation pattern might indicate the presence of different scrapie strains. Furthermore scrapie samples from goats in the current study demonstrated less long term resistance to proteinase K than ovine or caprine BSE control samples. Reduced scrapie susceptibility according to the PRNP genotype was demonstrated (Fishers Exact test, p < 0.05) for the goats with at least one polymorphism (p = 0.023) at the six codons examined and in particular for those with polymorphisms at codon 146 (p = 0.016). This work characterizes scrapie in goats having implications for breeding and surveillance strategies.
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Two Cheviot ewes homozygous for the A136L141R154Q171 (AL141RQ) prion protein (PrP) genotype were exposed intracerebrally to brain pools prepared using four field cases of atypical scrapie from the United Kingdom. Animals were clinically normal until the end of the experiment, when they were culled 7 years post-inoculation. Limited accumulation of disease-associated PrP (PrPSc) was observed in the cerebellar molecular layer by immunohistochemistry, but not by western blot or enzyme-linked immunosorbent assay. In addition, PrPSc was partially localized in astrocytes and microglia, suggesting that these cells have a role in PrPSc processing, degradation or both. Our results indicate that atypical scrapie is transmissible to AL141RQ sheep, but these animals act as clinically silent carriers with long incubation times.
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A clear association of amino acid variation in the prion protein gene (PRNP) with susceptibility and resistance to classical scrapie exists in sheep, but not in goats. In this study we examined DNA sequence variation in the PRNP of 149 animals from two scrapie-infected herds of Saanen dairy goats, and identified 6 non-synonymous variants in the coding region. In the larger herd, all of the 54 scrapie-affected goats tested had at least one allele with the arginine (R) codon at position 211, with 52 being homozygous for that variant. No animal homozygous for the glutamine (Q) codon at 211 were affected and only two heterozygotes (R/Q) were affected. A weak association was found at position 146 and no significant associations were found with amino acid variation at the remaining four variant positions (142, 143, 222 and 240), however, the allelic variation was low. Similar patterns were observed in the second scrapie-affected herd. We also evaluated previous studies on goat herds affected with scrapie and this relationship of R susceptibility and Q resistance at 211 was present independent of the genotypes at the other positions including 222. The fact that glutamine at 211 provides a significant protective property to scrapie irrespective of the other positions could be important for breeding strategies aimed at improving herd resistance to scrapie, while maintaining important productivity traits.
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The prion protein gene (Prnp) is highly influential in determining risk and susceptibility of sheep exposed to classical scrapie. Sheep homozygous for alanine at codon 136 and arginine at codons 154 and 171 (ARR/ARR) of the Prnp gene are historically considered to be highly resistant to classical scrapie, although they form a significant fraction of cases of atypical scrapie. To date, experimental transmission of prions to ARR/ARR sheep has only been achieved with the BSE agent and mostly by the intracerebral route. We summarise here the results of six separate studies, in which 95 sheep of the ARR/ARR genotype were naturally exposed to (n = 18) or experimentally challenged with (n = 77) natural or experimental sources of classical scrapie by the oral, intra-intestinal, subcutaneous or intracerebral routes and allowed to survive for periods of up to 94 months post-infection. Only the intracerebral route resulted in disease and/or amplification of disease associated PrP (PrPd), and only in two of 19 sheep that survived for longer than 36 months. Discriminatory immunohistochemistry and Western blot confirmed the scrapie, non-BSE signature of PrPd in those two sheep. However, the neuropathological phenotype was different from any other scrapie (classical or atypical) or BSE source previously reported in sheep of any Prnp genotype. These studies confirm the widely held view that ARR/ARR sheep are highly resistant to classical scrapie infection, at least within their commercial lifespan. Moreover, within the constraints of the present studies (only two infected sheep), these results do not support the suggestion that atypical scrapie or BSE are generated by adaptation or mutation of classical scrapie in sheep of resistant ARR/ARR genotype.
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Classical scrapie is one of the transmissible spongiform encephalopathies (TSEs), a group of fatal infectious diseases that affect the central nervous system (CNS). Classical scrapie can transmit laterally from ewe to lamb perinatally or between adult animals. Here we report detection of infectivity in tissues of an unborn fetus, providing evidence that in utero transmission of classical scrapie is also possible.
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In goats, several field studies have identified coding mutations of the gene encoding the prion protein (I/M142, N/D146, S/D146, R/Q211, and Q/K222) that are associated with a lower risk of developing classical scrapie. However, the data related to the levels of resistance to transmissible spongiform encephalopathies (TSEs) of these different PRNP gene mutations are still considered insufficient for developing large-scale genetic selection against scrapie in this species. In this study, we inoculated wild-type (WT) PRNP (I142R154R211Q222) goats and homozygous and/or heterozygous I/M142, R/H154, R/Q211, and Q/K222 goats with a goat natural scrapie isolate by either the oral or the intracerebral (i.c.) route. Our results indicate that the I/M142 PRNP polymorphism does not provide substantial resistance to scrapie infection following intracerebral or oral inoculation. They also demonstrate that H154, Q211, and K222 PRNP allele carriers are all resistant to scrapie infection following oral exposure. However, in comparison to WT animals, the H154 and Q211 allele carriers displayed only moderate increases in the incubation period following i.c. challenge. After i.c. challenge, heterozygous K222 and a small proportion of homozygous K222 goats also developed the disease, but with incubation periods that were 4 to 5 times longer than those in WT animals. These results support the contention that the K222 goat prion protein variant provides a strong but not absolutely protective effect against classical scrapie.
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Public and animal health controls to limit human exposure to animal prions are focused on bovine spongiform encephalopathy (BSE), but other prion strains in ruminants may also have zoonotic potential. One example is atypical/Nor98 scrapie, which evaded statutory diagnostic methods worldwide until the early 2000s. To investigate whether sheep infected with scrapie prions could be another source of infection, we inoculated transgenic mice that overexpressed human prion protein with brain tissue from sheep with natural field cases of classical and atypical scrapie, sheep with experimental BSE, and cattle with BSE. We found that these mice were susceptible to BSE prions, but disease did not develop after prolonged postinoculation periods when mice were inoculated with classical or atypical scrapie prions. These data are consistent with the conclusion that prion disease is less likely to develop in humans after exposure to naturally occurring prions of sheep than after exposure to epizootic BSE prions of ruminants.
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Prions are considered the best example to prove that the biological information can be transferred protein to protein through a conformational change. The term "prion-like" is used to describe molecular mechanisms that share similarities with the mammalian prion protein self-perpetuating aggregation and spreading characteristics. Since prions are presumably composed only of protein and are infectious, the more similar the mechanisms that occur in the different neurodegenerative diseases, the more these processes will resemble an infection. In vitro and in vivo experiments carried out during the last decade in different neurodegenerative disorders such as Alzheimer's disease (AD), Parkinson's diseases (PD), and amyotrophic lateral sclerosis (ALS) have shown a convergence toward a unique mechanism of misfolded protein propagation. In spite of the term "infection" that could be used to explain the mechanism governing the diversity of the pathological processes, other concepts as "seeding" or "de novo induction" are being used to describe the in vivo propagation and transmissibility of misfolded proteins. The current studies are demanding an extended definition of "disease-causing agents" to include those already accepted as well as other misfolded proteins. In this new scenario, "seeding" would be a type of mechanism by which an infectious agent can be transmitted but should not be used to define a whole "infection" process.
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A number of disease-associated PrP forms characterized by abnormally short proteinase K-resistant fragments (atypical PrPres) were recently described in prion diseases. The relationship between atypical PrPres and PrP(Sc), and their role in etiology of prion diseases, remains unknown. We examined the relationship between PrP(Sc) and atypical PrPres, a form characterized by short C-terminal proteinase K-resistant fragments, in a prion strain of synthetic origin. We found that the two forms exhibit distinct neuronal tropism, deposition patterns, and degree of pathological lesions. Immunostaining of brain regions demonstrated a partial overlap in anatomic involvement of the two forms and revealed the sites of their selective deposition. The experiments on amplification in vitro suggested that distinct neuronal tropism is attributed to differences in replication requirements, such as preferences for different cellular cofactors and PrP(C) glycoforms. Remarkably, deposition of atypical PrPres alone was not associated with notable pathological lesions, suggesting that it was not neurotoxic, but yet transmissible. Unlike PrP(Sc), atypical PrPres did not show significant perineuronal, vascular, or perivascular immunoreactivity. However, both forms showed substantial synaptic immunoreactivity. Considering that atypical PrPres is not associated with substantial lesions, this result suggests that not all synaptic disease-related PrP states are neurotoxic. The current work provides important new insight into our understanding of the structure-pathogenicity relationships of transmissible PrP states.
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Transmissible Spongiform Encephalopathies (TSEs), including scrapie in sheep, chronic wasting disease (CWD) in cervids, transmissible mink encephalopathy (TME), and bovine spongiform encephalopathy (BSE), are fatal diseases of the nervous system associated with accumulation of misfolded prion protein (PrPSc). Different strains of TSEs exist, associated with different PrPSc conformations that can be probed by the stability assay, in which PrPSc is treated with increasing concentrations of the denaturant guanidine hydrochloride (GdnHCl). Here, we provide the first comprehensive application of a rapid, protease-free version of the GdnHCl stability assay to brain tissue from cattle experimentally infected with various TSE isolates. Consistent with previous findings from a single Japanese isolate, the L-type isolates of BSE are not distinguishable from classical BSE in this assay. In contrast, H-type isolates of BSE, including our unique isolate of E211K BSE, exhibit higher stability than classical BSE, suggesting that its increased protection against protease digestion at the BSE N-terminus is associated with a higher stability in GdnHCl. While the difference in stability in our version of the assay is likely not large enough for effective use in a diagnostic laboratory setting, the use of alternative experimental conditions may enhance this effect. TSEs from other natural host species that have been passaged in cattle, including CWD and TME, were not distinguishable from classical BSE, while isolates of cattle passaged scrapie exhibited a slight increase in stability as compared to classical BSE. These results suggest that the core of PrPSc, as probed in this assay, has similar stability properties among cattle-passaged TSE isolates and that the conformational differences that lead to changes in the proteinase K cleavage site do not cause large changes in the stability of PrPSc from TSE-affected cattle. However, the stability differences observed here will provide a basis of comparison for new isolates of atypical BSE observed in the future and in other geographic locations, especially in the case of H-type BSE.
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Classical scrapie is a neurological disorder of the central nervous system (CNS) characterized by the accumulation of an abnormal, partially protease resistant prion protein (PrP(sc)) in the CNS and in some peripheral tissues in domestic small ruminants. Whereas the pathological changes and genetic susceptibility of ovine scrapie are well known, caprine scrapie has been less well studied. We report here a pathological study of 13 scrapie-affected goats diagnosed in Spain during the last 9 years. We used immunohistochemical and biochemical techniques to discriminate between classical and atypical scrapie and bovine spongiform encephalopathy (BSE). All the animals displayed PrP(sc) distribution patterns and western blot characteristics compatible with classical scrapie. In addition, we determined the complete open reading frame sequence of the PRNP in these scrapie-affected animals. The polymorphisms observed were compared with those of the herd mates (n = 665) and with the frequencies of healthy herds (n = 581) of native Spanish goats (Retinta, Pirenaica and Moncaina) and other worldwide breeds reared in Spain (Saanen, Alpine and crossbreed). In total, sixteen polymorphic sites were identified, including the known amino acid substitutions at codons G37V, G127S, M137I, I142M, H143R, R151H, R154H, R211Q, Q222K, G232W, and P240S, and new polymorphisms at codons G74D, M112T, R139S, L141F and Q215R. In addition, the known 42, 138 and 179 silent mutations were detected, and one new one is reported at codon 122. The genetic differences observed in the population studied have been attributed to breed and most of the novel polymorphic codons show frequencies lower than 5%. This work provides the first basis of polymorphic distribution of PRNP in native and worldwide goat breeds reared in Spain.
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The PrP gene polymorphisms at codons 142 (I/M), 154 (R/H), 211 (R/Q), 222 (Q/K) and 240 (S/P) and their association with susceptibility to classical scrapie infection were investigated in five French goat herds displaying a high disease prevalence (>10%). On the basis of PrPSc detection in central nervous system and in various lymphoid tissues, 301 out of the 1343 goats were found to be Scrapie infected. The statistical analyses indicated that while P240 mutation had no direct impact on scrapie infection risk, the H154, Q211 and K222 mutations were associated with high resistance to Scrapie. The M142 mutated allele was associated to a limited protection level against the disease. These results further reinforce the view that, like in sheep, the selection of certain PrP alleles could be envisaged to control and eradicate classical Scrapie in commercial goat population.
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It has long been established that the sheep Prnp genotype influences the susceptibility to scrapie, and some studies suggest that it can also determine several aspects of the disease phenotype. Other studies, however, indicate that the source of infection may also play a role in such phenotype. To address this question an experiment was set up in which either of two different natural scrapie sources, AAS from AA136 Suffolk and VVC from VV136 Cheviot sheep, were inoculated into AA136, VA136 and VV136 sheep recipients (n = 52). The immunohistochemical (IHC) profile of disease-associated PrP (PrPd) accumulation in the brain of recipient sheep was highly consistent upon codon 136 homologous and semi-homologous transmission, but could be either similar to or different from those of the inoculum donors. In contrast, the IHC profiles were highly variable upon heterologous transmission (VVC to AA136 and AAS to VV136). Furthermore, sheep of the same Prnp genotype could exhibit different survival times and PrPd profiles depending on the source of infection, and a correlation was observed between IHC and Western blot profiles. It was found that additional polymorphisms at codons 112 or 141 of AA136 recipients resulted in a delayed appearance of clinical disease or even in protection from infection. The results of this study strongly suggest that the scrapie phenotype in sheep results from a complex interaction between source, donor and recipient factors, and that the Prnp genotype of the recipient sheep does not explain the variability observed upon codon 136 heterologous transmissions, arguing for other genetic factors to be involved.
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Sheep scrapie susceptibility or resistance is a function of genotype, with polymorphisms at codon 171 in the sheep prion gene playing a major role. Glutamine (Q) at codon 171 contributes to scrapie susceptibility, while arginine (R) is associated with resistance. In some breeds, lysine (K) occurs at codon 171, but its effect on scrapie resistance has not been determined. Charge and structural similarities between K and R suggest that they may contribute to prion disease susceptibility in a similar way, but studies have not been performed to confirm this. The purpose of the current study was to compare susceptibility and incubation times of AA(136)RR(154)QQ(171) (where the letter denotes the amino acid and the number the position) with AA(136)RR(154)QK(171) sheep after inoculation with scrapie. Barbado AA(136)RR(154)QQ(171) and AA(136)RR(154)QK(171) sheep were inoculated with scrapie intracerebrally to assess their susceptibility to scrapie. After inoculation, sheep were observed daily for clinical signs and were euthanized and necropsied after clinical signs were unequivocal. Tissues were collected at necropsy for immunohistochemistry and Western blot analyses. The QQ(171) sheep had clinical signs approximately 12 months after inoculation, whereas QK(171) animals had an average incubation time of 30 months to onset of clinical signs. The distribution of abnormal prion protein was similar in QQ(171) and QK(171) sheep. Results of the study indicate that sheep with a single K allele at codon 171 are susceptible to scrapie but with a prolonged incubation time. Work is currently underway to examine relative scrapie susceptibility or resistance of KK(171) sheep.
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Scrapie of sheep and chronic wasting disease (CWD) of cervids are transmissible prion diseases. Milk and placenta have been identified as sources of scrapie prions but do not explain horizontal transmission. In contrast, CWD prions have been reported in saliva, urine and feces, which are thought to be responsible for horizontal transmission. While the titers of CWD prions have been measured in feces, levels in saliva or urine are unknown. Because sheep produce ~17 L/day of saliva, and scrapie prions are present in tongue and salivary glands of infected sheep, we asked if scrapie prions are shed in saliva. We inoculated transgenic (Tg) mice expressing ovine prion protein, Tg(OvPrP) mice, with saliva from seven Cheviot sheep with scrapie. Six of seven samples transmitted prions to Tg(OvPrP) mice with titers of -0.5 to 1.7 log ID₅₀ U/ml. Similarly, inoculation of saliva samples from two mule deer with CWD transmitted prions to Tg(ElkPrP) mice with titers of -1.1 to -0.4 log ID₅₀ U/ml. Assuming similar shedding kinetics for salivary prions as those for fecal prions of deer, we estimated the secreted salivary prion dose over a 10-mo period to be as high as 8.4 log ID₅₀ units for sheep and 7.0 log ID₅₀ units for deer. These estimates are similar to 7.9 log ID₅₀ units of fecal CWD prions for deer. Because saliva is mostly swallowed, salivary prions may reinfect tissues of the gastrointestinal tract and contribute to fecal prion shedding. Salivary prions shed into the environment provide an additional mechanism for horizontal prion transmission.
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Many prion diseases are orally acquired. Our data show that after oral exposure, early prion replication upon follicular dendritic cells (FDC) in Peyer's patches is obligatory for the efficient spread of disease to the brain (termed neuroinvasion). For prions to replicate on FDC within Peyer's patches after ingestion of a contaminated meal, they must first cross the gut epithelium. However, the mechanism through which prions are conveyed into Peyer's patches is uncertain. Within the follicle-associated epithelium overlying Peyer's patches are microfold cells (M cells), unique epithelial cells specialized for the transcytosis of particles. We show that following M cell-depletion, early prion accumulation upon FDC in Peyer's patches is blocked. Furthermore, in the absence of M cells at the time of oral exposure, neuroinvasion and disease development are likewise blocked. These data suggest M cells are important sites of prion uptake from the gut lumen into Peyer's patches.