Prion Disease In Cattle Research Articles

Sequence Analysis of Prion Protein Gene in Bhagnari Cattle Breed from the Hottest Region of Sibi, Balochistan, Pakistan

Prion diseases are a type of neurodegenerative disorder caused by the transmission of specific pathogens containing prion proteins. Due to the unique structural characteristics of Prion proteins (PrP), which differ from other types of proteins, the extended incubation period observed in the transmission of specific Prions can be attributed to these differences, at least in part. Prions are found in several other mammals and animals. Prions are unique among false protein folding abnormalities because these are infections and contain various strains of contagious agents associated with a unique in vivo phenotype. They can be acquired either by inheritance or sporadically. There are two types: classical and typical BSP. The objective of this study was to observe the Bhagnari cattle breed of Balochistan, Pakistan, and compare the PrP gene sequence of the Bhagnari with other reported sequences from Pakistan and other parts of the world. This research collected 40 Bhagnari cattle blood samples from Tali, Bhag Nari, and Sibi district areas. DNA extraction of each sample was performed by inorganic method, and then DNA amplification and sequencing of PRNP Gene was performed. The results of this research work showed different polymorphic variations (SNPs) in 16 samples. In this study, while mutations causing prion diseases in cattle were detected in Italian and German breeds, none were identified in the PRNP gene of the cattle population investigated, despite its association with neurological disorders.

The ultrastructure of infectious L-type bovine spongiform encephalopathy prions constrains molecular models.

Bovine spongiform encephalopathy (BSE) is a prion disease of cattle that is caused by the misfolding of the cellular prion protein (PrPC) into an infectious conformation (PrPSc). PrPC is a predominantly α-helical membrane protein that misfolds into a β-sheet rich, infectious state, which has a high propensity to self-assemble into amyloid fibrils. Three strains of BSE prions can cause prion disease in cattle, including classical BSE (C-type) and two atypical strains, named L-type and H-type BSE. To date, there is no detailed information available about the structure of any of the infectious BSE prion strains. In this study, we purified L-type BSE prions from transgenic mouse brains and investigated their biochemical and ultrastructural characteristics using electron microscopy, image processing, and immunogold labeling techniques. By using phosphotungstate anions (PTA) to precipitate PrPSc combined with sucrose gradient centrifugation, a high yield of proteinase K-resistant BSE amyloid fibrils was obtained. A morphological examination using electron microscopy, two-dimensional class averages, and three-dimensional reconstructions revealed two structural classes of L-type BSE amyloid fibrils; fibrils that consisted of two protofilaments with a central gap and an average width of 22.5 nm and one-protofilament fibrils that were 10.6 nm wide. The one-protofilament fibrils were found to be more abundant compared to the thicker two-protofilament fibrils. Both fibrillar assemblies were successfully decorated with monoclonal antibodies against N- and C-terminal epitopes of PrP using immunogold-labeling techniques, confirming the presence of polypeptides that span residues 100-110 to 227-237. The fact that the one-protofilament fibrils contain both N- and C-terminal PrP epitopes constrains molecular models for the structure of the infectious conformer in favour of a compact four-rung β-solenoid fold.

Experimental Study Using Multiple Strains of Prion Disease in Cattle Reveals an Inverse Relationship between Incubation Time and Misfolded Prion Accumulation, Neuroinflammation, and Autophagy

Proteinopathies result from aberrant folding and accumulation of specific proteins. Currently, there is a lack of knowledge about the factors that influence disease progression, making this a key challenge for the development of therapies for proteinopathies. Because of the similarities between transmissible spongiform encephalopathies (TSEs) and other protein misfolding diseases, TSEs can be used to understand other proteinopathies. Bovine spongiform encephalopathy (BSE) is a TSE that occurs in cattle and can be subdivided into three strains: classic BSE and atypical BSEs (H and L types) that have shorter incubation periods. The NACHT, LRR, and PYD domains-containing protein 3 inflammasome is a critical component of the innate immune system that leads to release of IL-1β. Macroautophagy is an intracellular mechanism that plays an essential role in protein clearance. In this study, the retina was used as a model to investigate the relationship between disease incubation period, prion protein accumulation, neuroinflammation, and changes in macroautophagy. We demonstrate that atypical BSEs present with increased prion protein accumulation, neuroinflammation, and decreased autophagy. This work suggests a relationship between disease time course, neuroinflammation, and the autophagic stress response, and may help identify novel therapeutic biomarkers that can delay or prevent the progression of proteinopathies.

Influence of Interspecies Transmission of Atypical Bovine Spongiform Encephalopathy Prions to Hamsters on Prion Characteristics.

Bovine spongiform encephalopathy (BSE) is a prion disease in cattle and is classified into the classical type (C-BSE) and two atypical BSEs, designated as high type (H-BSE) and low type (L-BSE). These classifications are based on the electrophoretic migration of the proteinase K-resistant core (PrPres) of the disease-associated form of the prion protein (PrPd). In a previous study, we succeeded in transmitting the H-BSE prion from cattle to TgHaNSE mice overexpressing normal hamster cellular PrP (PrPC). Further, Western blot analysis demonstrated that PrPres banding patterns of the H-BSE prion were indistinguishable from those of the C-BSE prion in TgHaNSE mice. In addition, similar PrPres glycoprofiles were detected among H-, C-, and L-BSE prions in TgHaNSE mice. Therefore, to better understand atypical BSE prions after interspecies transmission, H-BSE prion transmission from TgHaNSE mice to hamsters was investigated, and the characteristics of classical and atypical BSE prions among hamsters, wild-type mice, and mice overexpressing bovine PrPC (TgBoPrP) were compared in this study using biochemical and neuropathological methods. Identical PrPres banding patterns were confirmed between TgHaNSE mice and hamsters in the case of all three BSE prion strains. However, these PrPres banding patterns differed from those of TgBoPrP and wild-type mice infected with the H-BSE prion. In addition, glycoprofiles of TgHaNSE mice and hamsters infected with the L-BSE prion differed from those of TgBoPrP mice infected with the L-BSE prion. These data indicate that the PrPC amino acid sequences of new host species rather than other host environmental factors may affect some molecular aspects of atypical BSE prions. Although three BSE prion strains were distinguishable based on the neuropathological features in hamsters, interspecies transmission modified some molecular properties of atypical BSE prions, and these properties were indistinguishable from those of C-BSE prions in hamsters. Taken together, PrPres banding patterns and glycoprofiles are considered to be key factors for BSE strain typing. However, this study also revealed that interspecies transmission could sometimes influence these characteristics.

Detection of PrPBSE and prion infectivity in the ileal Peyer\u2019s patch of young calves as early as 2\xa0months after oral challenge with classical bovine spongiform encephalopathy

In classical bovine spongiform encephalopathy (C-BSE), an orally acquired prion disease of cattle, the ileal Peyer’s patch (IPP) represents the main entry port for the BSE agent. In earlier C-BSE pathogenesis studies, cattle at 4–6 months of age were orally challenged, while there are strong indications that the risk of infection is highest in young animals. In the present study, unweaned calves aged 4–6 weeks were orally challenged to determine the earliest time point at which newly formed PrPBSE and BSE infectivity are detectable in the IPP. For this purpose, calves were culled 1 week as well as 2, 4, 6 and 8 months post-infection (mpi) and IPPs were examined for BSE infectivity using a bovine PrP transgenic mouse bioassay, and for PrPBSE by immunohistochemistry (IHC) and protein misfolding cyclic amplification (PMCA) assays. For the first time, BSE prions were detected in the IPP as early as 2 mpi by transgenic mouse bioassay and PMCA and 4 mpi by IHC in the follicular dendritic cells (FDCs) of the IPP follicles. These data indicate that BSE prions propagate in the IPP of unweaned calves within 2 months of oral uptake of the agent.

Assessing the time taken for a surveillance system to detect a re-emergence of bovine spongiform encephalopathy in cattle

During the bovine spongiform encephalopathy (BSE) epidemic in July 2001 the European Commission established a surveillance scheme for the comprehensive sampling of all BSE clinical suspects, healthy slaughter (HS) animals >30months, and all emergency slaughter and fallen stock animals tested when >24months. With the exponential decline in classical BSE cases, this comprehensive surveillance system has been successively modified to become risk-based, targeting those exit streams and ages where cases from the original epidemic are most likely to be detected. Such reductions in testing are not without losses in the information subsequently collected, which could affect the sensitivity of the surveillance system to relatively small changes in the underlying prevalence of BSE across the European Union (EU). Here we report on a cohort-based approach to estimate the time taken for EU surveillance to observe a theoretical re-emergence of BSE in cattle. A number of surveillance schemes were compared. The baseline scheme considered detection being triggered by at least one case in the ‘age window’ 48–72 months in the fallen stock or emergency slaughter exit streams. Alternative schemes changed the start and end of this age window as well as considering testing for HS cattle. Under the baseline scheme, an estimated 15 years would lapse ([2.5th, 97.5th] percentiles=[10,24]) prior to detection, during which time 2867 infected animals ([2.5th, 97.5th]=[1722,6967]) would enter the slaughter population. These animals would be predominantly young animals (majority <24months) showing no clinical signs. This baseline scheme reduced the time to detection by 2 years, compared to a scheme where only clinical suspects were tested assuming BSE symptoms are recognised to the same degree by veterinary surgeons. Additional testing of younger animals did not improve detection as young infected animals were unlikely to test positive, but testing of older animals reduced the time to detection. Testing of HS animals >72months reduced the time to detection by one year compared to the baseline model, but would incur a high financial cost, e.g. testing HS animals >72months of age for 14 years would entail approximately 50.4 million additional tests. A limitation of the results is that there is no guarantee that current detection methods, optimised for detection of classical BSE, would identify a novel prion disease in cattle and it is currently difficult to envisage plausible routes by which a re-emergence of classical BSE could occur in Europe.

In vitro amplification of H-type atypical bovine spongiform encephalopathy by protein misfolding cyclic amplification

ABSTRACTThe in vitro amplification of prions by serial protein misfolding cyclic amplification has been shown to detect PrPSc to levels at least as sensitive as rodent bioassay but in a fraction of the time. Bovine spongiform encephalopathy is a zoonotic prion disease in cattle and has been shown to occur in 3 distinct forms, classical BSE (C-BSE) and 2 atypical BSE forms (L-BSE and H-BSE). Atypical forms are usually detected in asymptomatic, older cattle and are suggested to be spontaneous forms of the disease. Here, we show the development of a serial protein misfolding cyclic amplification method for the detection of H-BSE. The assay could detect PrPSc from 3 distinct experimental isolates of H-BSE, could detect PrPSc in as little as 1×10−12 g of brain material and was highly specific. Additionally, the product of serial protein misfolding cyclic amplification at all dilutions of seed analyzed could be readily distinguished from L-BSE, which did not amplify, and C-BSE, which had PrPSc with distinct protease K-resistance and protease K-resistant PrPSc molecular weights.

The Canadian Management of Bovine Spongiform Encephalopathy in Historical and Scientific Perspective, 1990-2014.

On February 11, 2015, the Canadian Food Inspection Agency announced that a cow born and raised in Alberta had tested positive for bovine spongiform encephalopathy (BSE), commonly known as mad cow disease. BSE is a prion disease of cattle that, when transmitted to humans, produces a fatal neurodegenerative disease known as variant Creutzfeldt-Jakob disease. We believe that this latest case of BSE in Canadian cattle suggests the timeliness of a review of the management of BSE in Canada from a historically and scientifically informed perspective. In this article, we ask: how did the Canadian management of BSE between 1990 and 2014 engage with the contemporary understanding of BSE's human health implications? We propose that Canadian policies largely ignored the implicit medical nature of BSE, treating it as a purely agricultural and veterinary issue. In this way, policies to protect Canadians were often delayed and incomplete, in a manner disturbingly reminiscent of Britain's failed management of BSE. Despite assurances to the contrary, it is premature to conclude that BSE (and with it the risk of variant Creutzfeldt-Jakob disease) is a thing of Canada's past: BSE remains very much an issue in Canada's present.

Genetics of Prion Disease in Cattle.

Bovine spongiform encephalopathy (BSE) is a prion disease that is invariably fatal in cattle and has been implicated as a significant human health risk. As a transmissible disease of livestock, it has impacted food safety, production practices, global trade, and profitability. Genetic polymorphisms that alter the prion protein in humans and sheep are associated with transmissible spongiform encephalopathy susceptibility or resistance. In contrast, there is no strong evidence that nonsynonymous mutations in the bovine prion gene (PRNP) are associated with classical BSE (C-BSE) disease susceptibility, though two bovine PRNP insertion/deletion polymorphisms, in the putative region, are associated with susceptibility to C-BSE. However, these associations do not explain the full extent of BSE susceptibility, and loci outside of PRNP appear to be associated with disease incidence in some cattle populations. This article provides a review of the current state of genetic knowledge regarding prion diseases in cattle.

Unique Properties of the Classical Bovine Spongiform Encephalopathy Strain and Its Emergence From H-Type Bovine Spongiform Encephalopathy Substantiated by VM Transmission Studies

In addition to classical bovine spongiform encephalopathy (C-BSE), which is recognized as being at the origin of the human variant form of Creutzfeldt-Jakob disease, 2 rare phenotypes of BSE (H-type BSE [H-BSE] and L-type BSE [L-BSE]) were identified in 2004. H-type BSE and L-BSE are considered to be sporadic forms of prion disease in cattle because they differ from C-BSE with respect to incubation period, vacuolar pathology in the brain, and biochemical properties of the protease-resistant prion protein (PrP) in natural hosts and in some mouse models that have been tested. Recently, we showed that H-BSE transmitted to C57Bl/6 mice resulted in a dissociation of the phenotypic features, that is, some mice showed an H-BSE phenotype, whereas others had a C-BSE phenotype. Here, these 2 phenotypes were further studied in VM mice and compared with cattle C-BSE, H-BSE, and L-BSE. Serial passages from the C-BSE-like phenotype on VM mice retained similarities with C-BSE. Moreover, our results indicate that strains 301V and 301C derived from C-BSE transmitted to VM and C57Bl/6 mice, respectively, are fundamentally the same strain. These VM transmission studies confirm the unique properties of the C-BSE strain and support the emergence of a strain that resembles C-BSE from H-BSE.

Human prion diseases: Molecular, cellular and population biology

The past 20 years have witnessed a dramatic resurgence of interest in a hitherto obscure neurodegenerative disease, Creutzfeldt‐Jakob disease (CJD). This was driven partly by the novelty of the prion hypothesis, which sought to provide an explanation for the pathogenesis of transmissible spongiform encephalopathies, involving a unique epigenetic mechanism, and partly by events in the UK, where an outbreak of a new prion disease in cattle (bovine spongiform encephalopathy or BSE) potentially exposed a large section of the UK population to prion infectivity through a dietary route. The numbers of cases of the resultant novel disease variant CJD (vCJD), have so far been limited and peaked in the UK in the year 2000 and have subsequently declined. However, the effects of BSE and vCJD have been far‐reaching. The estimated prevalence of vCJD infection in the UK is substantially higher than the numbers of clinical cases would suggest, posing a difficult dilemma for those involved in blood transfusion, tissue transplantation and cellular therapies. The clinico‐pathological phenotype of human prion diseases has come under close scrutiny and molecular classification systems have been developed to account for the different diseases and their phenotypic spectra. Moreover, enhanced human and animal surveillance and better diagnostic tools have identified new human and animal prion diseases. Lastly, as the prion hypothesis has gained widespread acceptance, the concepts involved have been applied to other areas, including extra‐chromosomal inheritance in fungi, long‐term potentiation in memory formation and the spread of molecular pathology in diverse conditions, such as Alzheimer's disease, Parkinson's disease and amyotrophic lateral sclerosis. Studies at the molecular and cellular level have helped to provide a better understanding of human prion diseases, aided pathological diagnosis and helped inform public health decision‐making.

Four BSE cases with an L-BSE molecular profile in cattle from Great Britain

Bovine spongiform encephalopathy (BSE) is a prion disease of cattle which was first observed in Great Britain (GB) in 1986. Throughout the subsequent BSE epidemic, cases identified by passive surveillance have shown consistent histopathological, immunohistochemical, biochemical and biological properties. However, since the start of active surveillance in 2001, across Europe and elsewhere, approximately 67 cases with different biochemical characteristics have been identified by Western blotting (WB). These cases fall into two categories; 'H-type' (H-BSE) or 'L-type' (L-BSE), based on the relatively heavy (H-BSE) or light (L-BSE) mass of the unglycosylated band of the prion protein, as compared with WB against that obtained from classical BSE (C-BSE) cases. Here we report the detection and confirmation of the first four L-BSE cases by active surveillance in GB, two of which were born after the reinforced feed ban of 1996 (BARB cases). These four L-BSE cases were found in relatively old cattle (age range; 11-21 years old) and the carcases did not enter the human food chain or animal feed chains.

Four BSE cases with an L-BSE molecular profile in cattle from Great Britain

Bovine spongiform encephalopathy (BSE) is a prion disease of cattle which was first observed in Great Britain (GB) in 1986. Throughout the subsequent BSE epidemic, cases identified by passive surveillance...

Susceptibility of cattle to the agent of chronic wasting disease from elk after intracranial inoculation

Cattle could be exposed to the agent of chronic wasting disease (CWD) through contact with infected farmed or free-ranging cervids or exposure to contaminated premises. The purpose of the current study was to assess the potential for CWD derived from elk to transmit to cattle after intracranial inoculation. Calves (n = 14) were inoculated with brain homogenate derived from elk with CWD to determine the potential for transmission and to define the clinicopathologic features of disease. Cattle were necropsied if clinical signs occurred or at the end of the study (49 months postinoculation; MPI). Clinical signs of poor appetite, weight loss, circling, and bruxism occurred in 2 cattle (14%) at 16 and 17 MPI, respectively. Accumulation of abnormal prion protein (PrP(Sc)) occurred in only the 2 clinically affected cattle and was confined to the central nervous system, with the most prominent immunoreactivity in midbrain, brainstem, and hippocampus with lesser immunoreactivity in the cervical spinal cord. The rate of transmission was lower than in cattle inoculated with CWD derived from mule deer (38%) or white-tailed deer (86%). Additional studies are required to fully assess the potential for cattle to develop CWD through a more natural route of exposure, but a low rate of transmission after intracranial inoculation suggests that risk of transmission through other routes is low. A critical finding is that if CWD did transmit to exposed cattle, currently used diagnostic techniques would detect and differentiate it from other prion diseases in cattle based on absence of spongiform change, distinct pattern of PrP(Sc) deposition, and unique molecular profile.

High sensitivity detection of the glial fibrillary acidic protein as indicator for TSE risk material in meat products using an immuno‐PCR

The emergence of prion diseases in cattle during the bovine spongiform encephalopathy (BSE) epidemic and the transmission to humans causing variant Creutzfeldt-Jakob disease by consume of BSE-contaminated meat has focused attention on the use of tissues from the central nervous system (CNS) in food. To avoid food contamination, it is regulated by law that specified risk material has to be removed from food chains. Detection of well-expressed CNS indicator proteins such as the glial fibrillary acidic protein (GFAP) assumes an important role in detection of food contamination; however, the sensitivity of detection performed on basis of ELISAs is limited. Consequently, there is an urgent need for high-sensitivity detection methods. We describe the development of an immuno-PCR assay with enhancement of the immunoreaction by amplification of a DNA fragment linked to the antigen-antibody complex. This immuno-PCR assay shows enhanced sensitivity in GFAP detection of at least two orders of magnitude compared to the ELISA technique carried out under identical conditions. The immuno-PCR proves to be a suitable tool for the high-sensitivity detection of marker proteins in food imported by food chain processes. It is applicable for use by food and veterinary facilities, and institutes involved in food monitoring.

Evaluation of the Human Transmission Risk of an Atypical Bovine Spongiform Encephalopathy Prion Strain

Bovine spongiform encephalopathy (BSE), the prion disease in cattle, was widely believed to be caused by only one strain, BSE-C. BSE-C causes the fatal prion disease named new variant Creutzfeldt-Jacob disease in humans. Two atypical BSE strains, bovine amyloidotic spongiform encephalopathy (BASE, also named BSE-L) and BSE-H, have been discovered in several countries since 2004; their transmissibility and phenotypes in humans are unknown. We investigated the infectivity and human phenotype of BASE strains by inoculating transgenic (Tg) mice expressing the human prion protein with brain homogenates from two BASE strain-infected cattle. Sixty percent of the inoculated Tg mice became infected after 20 to 22 months of incubation, a transmission rate higher than those reported for BSE-C. A quarter of BASE strain-infected Tg mice, but none of the Tg mice infected with prions causing a sporadic human prion disease, showed the presence of pathogenic prion protein isoforms in the spleen, indicating that the BASE prion is intrinsically lymphotropic. The pathological prion protein isoforms in BASE strain-infected humanized Tg mouse brains are different from those from the original cattle BASE or sporadic human prion disease. Minimal brain spongiosis and long incubation times are observed for the BASE strain-infected Tg mice. These results suggest that in humans, the BASE strain is a more virulent BSE strain and likely lymphotropic.

Conversion of the BASE Prion Strain into the BSE Strain: The Origin of BSE?

Atypical neuropathological and molecular phenotypes of bovine spongiform encephalopathy (BSE) have recently been identified in different countries. One of these phenotypes, named bovine “amyloidotic” spongiform encephalopathy (BASE), differs from classical BSE for the occurrence of a distinct type of the disease-associated prion protein (PrP), termed PrPSc, and the presence of PrP amyloid plaques. Here, we show that the agents responsible for BSE and BASE possess different biological properties upon transmission to transgenic mice expressing bovine PrP and inbred lines of nontransgenic mice. Strikingly, serial passages of the BASE strain to nontransgenic mice induced a neuropathological and molecular disease phenotype indistinguishable from that of BSE-infected mice. The existence of more than one agent associated with prion disease in cattle and the ability of the BASE strain to convert into the BSE strain may have important implications with respect to the origin of BSE and spongiform encephalopathies in other species, including humans.

Prions in Deer Muscle

Prion diseases in cattle and sheep predominantly affect nervous system tissue. Chronic wasting disease in deer is a prion disease. Because humans consume deer muscle (meat), a multi-institutional team used a sensitive assay to look for prions in …

Chronic Wasting Disease of Elk: Transmissibility to Humans Examined by Transgenic Mouse Models

Chronic wasting disease (CWD), a prion disease affecting free-ranging and captive cervids (deer and elk), is widespread in the United States and parts of Canada. The large cervid population, the popularity of venison consumption, and the apparent spread of the CWD epidemic are likely resulting in increased human exposure to CWD in the United States. Whether CWD is transmissible to humans, as has been shown for bovine spongiform encephalopathy (the prion disease of cattle), is unknown. We generated transgenic mice expressing the elk or human prion protein (PrP) in a PrP-null background. After intracerebral inoculation with elk CWD prion, two lines of "humanized" transgenic mice that are susceptible to human prions failed to develop the hallmarks of prion diseases after >657 and >756 d, respectively, whereas the "cervidized" transgenic mice became infected after 118-142 d. These data indicate that there is a substantial species barrier for transmission of elk CWD to humans.

Diagnosis and Incidence of Prion (Creutzfeldt-Jakob) Disease: A Retrospective Archival Survey with Implications for Future Research

Reliable identification of Creutzfeldt-Jakob disease (CJD) in the UX has become essential following the suggestion that prion disease in cattle (BSE) might transmit, accidentally, to humans who eat contaminated beef. Recent data suggest that some cases of CJD may be clinically unrecognized; in order to examine this proposal we reviewed all cases of dementia ( n=1000+) collected in the Runwell Hospital Brain Archive between 1964 and 1990. We identified 19 cases of spongiform encephalopathy of which only 11 were diagnosed before death. These 11 individuals had a characteristic clinical history of CJD (relentless mental deterioration, prominent motor signs and death within a year). Their brains showed little or no external abnormality. In contrast, only two of the eight clinically unrecognized cases had characteristic symptoms. The remaining six presented atypically; their illness lasted 3 years or more, motor signs were much less evident, and simple dementia was the most prominent feature. The brains showed moderate or severe cerebral atrophy. Our data indicate that only about 60% of prion disease cases with pathologically typical spongiform encephalopathy were identified clinically during life. This suggests that human prion disease may be more common than previously supposed and that a further review of the epidemiology of the disease is required