Susceptibility of cattle to first-passage intracerebral inoculation with chronic wasting disease agent from white-tailed deer

Fourteen, 3-month-old calves were intracerebrally inoculated with the agent of chronic wasting disease (CWD) from white-tailed deer (CWDwtd) to compare the clinical signs and neuropathologic findings with those of certain other transmissible spongiform encephalopathies (TSE, prion diseases) that have been shown to be experimentally transmissible to cattle (sheep scrapie, CWD of mule deer [CWDmd], bovine spongiform encephalopathy [BSE], and transmissible mink encephalopathy). Two uninoculated calves served as controls. Within 26 months postinoculation (MPI), 12 inoculated calves had lost considerable weight and eventually became recumbent. Of the 12 inoculated calves, 11 (92%) developed clinical signs. Although spongiform encephalopathy (SE) was not observed, abnormal prion protein (PrPd) was detected by immunohistochemistry (IHC) and Western blot (WB) in central nervous system tissues. The absence of SE with presence of PrPd has also been observed when other TSE agents (scrapie and CWDmd) were similarly inoculated into cattle. The IHC and WB findings suggest that the diagnostic techniques currently used to confirm BSE would detect CWDwtd in cattle, should it occur naturally. Also, the absence of SE and a distinctive IHC pattern of CWDwtd and CWDmd in cattle suggests that it should be possible to distinguish these conditions from other TSEs that have been experimentally transmitted to cattle.     

Molecular profiling and comparison of field transmissible spongiform encephalopathy cases diagnosed in Catalunya

Molecular profiling of the proteinase K resistant prion protein (PrP(res)) is a technique that has been applied to the characterisation of transmissible spongiform encephalopathy (TSE) strains. An interesting example of the application of this technique is the ability to differentiate, at the experimental level, between bovine spongiform encephalopathy (BSE) and scrapie infection in sheep, and to distinguish between classical and atypical BSE and scrapie cases. Twenty-six BSE cases and two scrapie cases from an active TSE surveillance program and diagnosed at the PRIOCAT, TSE Reference Laboratory (Centre de Recerca en Sanitat Animal, Universitat Autònoma de Barcelona, Catalunya, Spain) were examined by Western blotting. Molecular profiling was achieved by comparing the glycosylation profile, deglycosylated PrP molecular weight and 6H4/P4 monoclonal antibody binding ratio. The results obtained during the characterisation of these field cases indicated an absence of atypical BSE cases in Catalunya.     

First and second cattle passage of transmissible mink encephalopathy by intracerebral inoculation

To compare clinicopathologic findings of transmissible mink encephalopathy (TME) with other transmissible spongiform encephalopathies (TSE, prion diseases) that have been shown to be experimentally transmissible to cattle (sheep scrapie and chronic wasting disease [CWD]), two groups of calves (n = 4 each) were intracerebrally inoculated with TME agents from two different sources (mink with TME and a steer with TME). Two uninoculated calves served as controls. Within 15.3 months postinoculation, all animals from both inoculated groups developed clinical signs of central nervous system (CNS) abnormality; their CNS tissues had microscopic spongiform encephalopathy (SE); and abnormal prion protein (PrP(res)) as detected in their CNS tissues by immunohistochemistry (IHC) and Western blot (WB) techniques. These findings demonstrate that intracerebrally inoculated cattle not only amplify TME PrP(res) but also develop clinical CNS signs and extensive lesions of SE. The latter has not been shown with other TSE agents (scrapie and CWD) similarly inoculated into cattle. The findings also suggest that the diagnostic techniques currently used for confirmation of bovine spongiform encephalopathy (BSE) would detect TME in cattle should it occur naturally. However, it would be a diagnostic challenge to differentiate TME in cattle from BSE by clinical signs, neuropathology, or the presence of PrP(res) by IHC and WB.     

PrP genetics in ruminant transmissible spongiform encephalopathies

Scrapie, bovine spongiform encephalopathy (BSE), and chronic wasting disease (CWD) are prion diseases in ruminants with considerable impact on animal health and welfare. They can also pose a risk to human health and control is therefore an important issue. Prion protein (PrP) genetics may be used to control and eventually eradicate animal prion diseases. The PrP gene in sheep and other representatives of the order Artiodactyles has many polymorphisms of which several are crucial determinants of susceptibility to prion diseases, also known as transmissible spongiform encephalopathies (TSE). This review will present the current understanding of PrP genetics in ruminants highlighting similarity and difference between the species in the context of TSE.     

Strain typing of German transmissible spongiform encephalopathies field cases in small ruminants by biochemical methods

Following the implementation of a large scale transmissible spongiform encephalopathies (TSE) surveillance programme of small ruminants, evidence for a natural transmission of bovine spongiform encephalopathy (BSE) to a French goat has been found. During the years 2002-2004, a massive TSE rapid testing programme on >250,000 small ruminants was carried out in Germany. In this national survey, 186 scrapie-affected sheep were found which originated from 78 flocks. The majority of these cases were of the classical TSE type (115 sheep belonging to 14 outbreaks). However, 71 cases coming from 64 flocks were of the novel atypical scrapie type. According to the regulation EU 999/2001, all TSE cases in small ruminants have to be examined by strain typing methods to explore any possibility of the existence of BSE cases in the field sheep population. Here we report on a biochemical typing strategy (termed FLI-test), which includes the determination of molecular masses, antibody binding affinities and glycosylation pattern of the TSE induced abnormal prion protein. Based on this typing approach none of the analysed German classical TSE outbreaks (total number of analysed sheep: 36) displayed biochemical features indicative for a BSE infection. However, in two cases distinct but BSE-unrelated PrP(Sc) types were found, which alludes to the existence of different scrapie strains in the German sheep population.     

Properties of L-type bovine spongiform encephalopathy in intraspecies passages

The origin and transmission routes of atypical bovine spongiform encephalopathy (BSE) remain unclear. To assess whether the biological and biochemical characteristics of atypical L-type BSE detected in Japanese cattle (BSE/JP24) are conserved during serial passages within a single host, 3 calves were inoculated intracerebrally with a brain homogenate prepared from first-passaged BSE/JP24-affected cattle. Detailed immunohistochemical and neuropathologic analysis of the brains of second-passaged animals, which had developed the disease and survived for an average of 16 months after inoculation, revealed distribution of spongiform changes and disease-associated prion protein (PrP(Sc)) throughout the brain. Although immunolabeled PrP(Sc) obtained from brain tissue was characterized by the presence of PrP plaques and diffuse synaptic granular accumulations, no stellate-type deposits were detected. Western blot analysis suggested no obvious differences in PrP(Sc) molecular mass or glycoform pattern in the brains of first- and second-passaged cattle. These findings suggest failures to identify differences in mean incubation period and biochemical and neuropathologic properties of the BSE/JP24 prion between the first and second passages in cattle.     

Atypical scrapie in a Swiss goat and implications for transmissible spongiform encephalopathy surveillance.

Different types of transmissible spongiform encephalopathies (TSEs) affect sheep and goats. In addition to the classical form of scrapie, both species are susceptible to experimental infections with the bovine spongiform encephalopathy (BSE) agent, and in recent years atypical scrapie cases have been reported in sheep from different European countries. Atypical scrapie in sheep is characterized by distinct histopathologic lesions and molecular characteristics of the abnormal scrapie prion protein (PrP(sc)). Characteristics of atypical scrapie have not yet been described in detail in goats. A goat presenting features of atypical scrapie was identified in Switzerland. Although there was no difference between the molecular characteristics of PrP(sc) in this animal and those of atypical scrapie in sheep, differences in the distribution of histopathologic lesions and PrP(sc) deposition were observed. In particular the cerebellar cortex, a major site of PrP(sc) deposition in atypical scrapie in sheep, was found to be virtually unaffected in this goat. In contrast, severe lesions and PrP(sc) deposition were detected in more rostral brain structures, such as thalamus and midbrain. Two TSE screening tests and PrP(sc) immunohistochemistry were either negative or barely positive when applied to cerebellum and obex tissues, the target samples for TSE surveillance in sheep and goats. These findings suggest that such cases may have been missed in the past and could be overlooked in the future if sampling and testing procedures are not adapted. The epidemiological and veterinary public health implications of these atypical cases, however, are not yet known.     

TSE pathogenesis in cattle and sheep

Many studies have been undertaken in rodents to study the pathogenesis of transmissible spongiform encephalopathies (TSE). Only a few studies have focused on the pathogenesis of bovine spongiform encephalopathy (BSE) and scrapie in their natural hosts. In this review, we summarize the most recent insights into the pathogenesis of BSE and scrapie starting from the initial uptake of TSE agents and crossing of the gut epithelium. Following replication in the gut-associated lymphoid tissues (GALT), TSE agents spread to the enteric nervous system (ENS) of the gut. Infection is then carried through the efferent fibers of the post-ganglionic neurons of the parasympathetic and sympathetic nervous system to the pre-ganglionic neurons in the medulla oblongata of the brain and the thoracic segments of the spinal cord. The differences between the pathogenesis of BSE in cattle and scrapie in sheep are discussed as well as the possible existence of additional pathogenetic routes.     

牛朊蛋白基因多态性对疯牛病影响的研究进展

朊蛋白(prion protein,PRNP)是近年来已证明的人和部分哺乳动物传染性海绵状脑病(transmissible spongiform encephalopathy,TSE)的主要根源,该蛋白编码基因的多态性显著影响了人和动物对TSE的易感性或抗病性。牛传染性海绵状脑病俗称"疯牛病"。作者分析了疯牛病的起源、监测和预防措施;简要介绍了牛PRNP基因的结构与功能;系统分析了牛科动物PRNP基因非编码区多态性与抗病性作用;总结了牛科动物PRNP基因启动子区域内23 bp插入/缺失和第1内含子区域内12 bp插入/缺失对疯牛病易感性的影响,为牛的抗病分子育种提供指导。     

Bovine spongiform encephalopathy in ruminants and the new variant of Creutzfeldt-Jakob disease in humans]

The feeding of meat and bone meal which was contaminated with bovine spongiform encephalopathy (BSE) infectivity led to a huge epidemic in the British cattle population. After the emergence of a new variant form of Creutzfeldt-Jakob diseases (nvCJD) in 1996, European consumers lost confidence in beef meat. Given the coincidence by time and place and disease and agent specific characteristics, it must be assumed that nvCJD is caused by the transmission of BSE agent to man. This article gives an overview over the characteristics and diagnosis of transmissible spongiform encephalopathies, namely BSE and nvCJD, in animals and man.     

Prion biology relevant to bovine spongiform encephalopathy

Bovine spongiform encephalopathy (BSE) and chronic wasting disease (CWD) of deer and elk are a threat to agriculture and natural resources, as well as a human health concern. Both diseases are transmissible spongiform encephalopathies (TSE), or prion diseases, caused by autocatalytic conversion of endogenously encoded prion protein (PrP) to an abnormal, neurotoxic conformation designated PrPsc. Most mammalian species are susceptible to TSE, which, despite a range of species-linked names, is caused by a single highly conserved protein, with no apparent normal function. In the simplest sense, TSE transmission can occur because PrPsc is resistant to both endogenous and environmental proteinases, although many details remain unclear. Questions about the transmission of TSE are central to practical issues such as livestock testing, access to international livestock markets, and wildlife management strategies, as well as intangible issues such as consumer confidence in the safety of the meat supply. The majority of BSE cases seem to have been transmitted by feed containing meat and bone meal from infected animals. In the United Kingdom, there was a dramatic decrease in BSE cases after neural tissue and, later, all ruminant tissues were banned from ruminant feed. However, probably because of heightened awareness and widespread testing, there is growing evidence that new variants of BSE are arising "spontaneously," suggesting ongoing surveillance will continue to find infected animals. Interspecies transmission is inefficient and depends on exposure, sequence homology, TSE donor strain, genetic polymorphism of the host, and architecture of the visceral nerves if exposure is by an oral route. Considering the low probability of interspecies transmission, the low efficiency of oral transmission, and the low prion levels in nonnervous tissues, consumption of conventional animal products represents minimal risk. However, detection of rare events is challenging, and TSE literature is characterized by subsequently unsupported claims of species barriers or absolute tissue safety. This review presents an overview of TSE and summarizes recent research on pathogenesis and transmission.     

Distribution of a pathological form of prion protein in the brainstem and cerebellum in classical and atypical cases of bovine spongiform encephalopathy

This study evaluated the distribution and signal intensity of a prion protein resistant to proteolysis (PrP(res)) in the brainstem and cerebellum of cattle affected with classical and atypical forms of bovine spongiform encephalopathy (BSE) using a Western immunoblotting technique. In both classical and atypical cases of BSE, a stronger signal was detected in the more rostral brainstem regions relative to the obex. In classical and H-type cases a significant decrease in the PrP(res) signal was found in the cerebellum when compared to that in the obex, whereas L-type BSE cases were characterised by signals of similar intensity in these regions. The uniform distribution of PrP(res) in the region rostral to the obex suggests that when autolysed samples are being tested for BSE, both classical and atypical forms are detectable, even when this target site is missing or cannot be clearly identified. The findings indicate that both the obex and rostral brainstem can be used for BSE diagnosis whereas use of the more caudal brainstem regions and cerebellum is not recommended.     

Approaches to investigating transmission of spongiform encephalopathies in domestic animals using BSE as an example

Bovine spongiform encephalopathy was a novel spongiform encephalopathy, in an hitherto unaffected species, that had characteristics of a point source epidemic, with an agent that could have been incorporated into a wide variety of feedstuffs and iatrogenically administered to na?ve populations, and there was early evidence that it was not restricted to bovines. It was vital to establish, albeit experimentally, which other species might be affected, and whether the epidemic could be maintained by natural transmission, if the source was removed. In contrast, scrapie has been endemic throughout Great Britain for centuries, is maintained naturally (even if we don't know exactly how) and has a known host range. The principles, process and integration of evidence from different types of studies, however, are similar for both of these transmissible spongiform encephalopathies (TSE) and can be applied to any emerging or suspected spongiform encephalopathy. This review discusses the experimental approaches used to determine TSE transmissibility and infectivity and how they relate to natural disease and control measures.     

Sequence variations of the bovine prion protein gene (PRNP) in native Korean Hanwoo cattle

Bovine spongiform encephalopathy (BSE) is one of the fatal neurodegenerative diseases known as transmissible spongiform encephalopathies (TSEs) caused by infectious prion proteins. Genetic variations correlated with susceptibility or resistance to TSE in humans and sheep have not been reported for bovine strains including those from Holstein, Jersey, and Japanese Black cattle. Here, we investigated bovine prion protein gene (PRNP) variations in Hanwoo cattle [Bos (B.) taurus coreanae], a native breed in Korea. We identified mutations and polymorphisms in the coding region of PRNP, determined their frequency, and evaluated their significance. We identified four synonymous polymorphisms and two non-synonymous mutations in PRNP, but found no novel polymorphisms. The sequence and number of octapeptide repeats were completely conserved, and the haplotype frequency of the coding region was similar to that of other B. taurus strains. When we examined the 23-bp and 12-bp insertion/deletion (indel) polymorphisms in the non-coding region of PRNP, Hanwoo cattle had a lower deletion allele and 23-bp del/12-bp del haplotype frequency than healthy and BSE-affected animals of other strains. Thus, Hanwoo are seemingly less susceptible to BSE than other strains due to the 23-bp and 12-bp indel polymorphisms.     

Beyond BSE: Transmissible spongiform encephalopathies in other animal species

There are several other diseases besides BSE which belong to the group of transmissible spongiform encephalopathies (TSE). Although most mammals can be experimentally infected with the agent of these diseases, generally only single representatives of the orders Artiodactyla (cloven-hoofed animals), Carnivora (carnivores) and Primates (humans and monkeys) are naturally infected in the field. An overview of the current state of knowledge on TSE in several species like exotic ruminants, deer, mink and cats is presented. Etiological, clinical, anatomic-pathological and epidemiological aspects are described.     

Bovine spongiform encephalopathy: detection and quantitation of fibrils, fibril protein (PrP) and vacuolation in brain

Bovine spongiform encephalopathy (BSE) is a new disease of cattle which has considerable homology with scrapie, the archetype of the transmissible spongiform encephalopathies. Abnormal brain fibrils, called scrapie associated fibrils (SAF), are specific ultrastructural markers for these diseases. Fibril detection was compared with histopathological diagnosis in the brains of 167 cattle; 157 clinically suspect BSE and 10 clinically normal. Fibrils were detected in samples of pooled brain regions of 67/144 in which vacuolar changes of BSE were confirmed, but absent in the remaining 23 brains, in which no vacuolation was found, including those from the clinically normal cattle and 13 with alternative neuropathological diagnoses. When eight defined anatomic regions from the brains of another 22 affected cows were examined, the sensitivity of fibril detection was greater than 90% for the brain stem areas. Fibril prevalence in these areas approximated to severity of vacuolar changes. When the same defined regions from four of the affected cows were assayed for fibril protein (PrP) by western blotting, the density of immuno-labelling generally correlated with the fibril prevalence. This study confirms the specificity of fibril detection for BSE, shows that the ease of fibril detection depends on anatomic region sampled and suggests an association between PrP accumulation and vacuolar changes in certain neuroanatomic areas.     

羊痒病研究进展

牛的海绵状脑病（BSE）的爆发让人们意识到要控制传染性海绵状脑病（TSE）的流行来保护人和动物的健康。羊痒病是世界上传播最广的传染性海绵状脑病,为此,不同国家的兽医当局都有针对痒病的根除计划,除了增强对动物的监管外,还要培育对痒病有抗性的羊。笔者主要综述了当前羊痒病的研究进展。     

Changes in plasma metabolites and muscle glycogen are correlated to bovine spongiform encephalopathy in infected dairy cattle

During the clinical phase of bovine spongiform encephalopathy (BSE), a significant decrease was observed in the ratio of muscle glycogen to plasma L-lactic acid concentrations in BSE infected field case and experimentally infected dairy cattle compared with healthy control cattle (P<0.001), this being due to changes in the concentration of both metabolites in the BSE infected cattle compared with the control group. Furthermore, the concentration of plasma alanine was significantly increased (P<0.05) in the infected animals. No significant difference was detected between these two groups in the ratio of hepatic glycogen to plasma lactate. We infer that BSE infected cattle exhibit signs of altered energy metabolism and when applied in conjunction with changes in other metabolite biomarkers these changes may be useful for discriminating BSE infected cattle from healthy cattle or those suffering with other disorders or diseases.     

Identification and characterization of two bovine spongiform encephalopathy cases diagnosed in the United States.

Bovine spongiform encephalopathy (BSE) is a transmissible spongiform encephalopathy of cattle, first detected in 1986 in the United Kingdom and subsequently in other countries. It is the most likely cause of variant Creutzfeldt-Jakob disease (vCJD) in humans, but the origin of BSE has not been elucidated so far. This report describes the identification and characterization of two cases of BSE diagnosed in the United States. Case 1 (December 2003) exhibited spongiform changes in the obex area of the brainstem and the presence of the abnormal form of the prion protein, PrP(Sc), in the same brain area, by immunohistochemistry (IHC) and Western blot analysis. Initial suspect diagnosis of BSE for case 2 (November 2004) was made by a rapid ELISA-based BSE test. Case 2 did not exhibit unambiguous spongiform changes in the obex area, but PrP(Sc) was detected by IHC and enrichment Western blot analysis in the obex. Using Western blot analysis, PrP(Sc) from case 1 showed molecular features similar to typical BSE isolates, whereas PrP(Sc) from case 2 revealed an unusual molecular PrP(Sc) pattern: molecular mass of the unglycosylated and monoglycosylated isoform was higher than that of typical BSE isolates and case 2 was strongly labeled with antibody P4, which is consistent with a higher molecular mass. Sequencing of the prion protein gene of both BSE-positive animals revealed that the sequences of both animals were within [corrected] the range of the prion protein gene sequence diversity previously reported for cattle.     

Risk assessment and surveillance for bovine spongiform encephalopathy (BSE) in Argentina

An assessment was made of the risks of bovine spongiform encephalopathy (BSE) occurring in Argentina. Most of the factors associated with the origin and development of the BSE epidemic in the UK are essentially absent. For example, Argentina's large sheep and cattle industries are based on low-cost production systems using grass. Concentrated feeds are not used for sheep, rarely for beef cattle and to a comparatively modest extent for dairy cows. Particularly important are the facts that scrapie (and BSE) has never been reported in Argentina—very small amounts of waste tissues from sheep are rendered to produce meat and bone meal (MBM)—and MBM is not used in concentrated feeds for cattle. We conclude that Argentina has an exceptionally low risk of BSE due to scrapie. There is a very small risk of BSE having been introduced via live animals imported from countries with BSE, but this could only give rise to isolated cases because MBM is not fed to cattle.

A surveillance programme has been carried out based largely on a histological examination of brains from three categories of old dairy cows: animals reported on the suspicion of having neurological disease; animals in poor condition at slaughter; healthy animals randomly selected in the abattoir. No evidence of transmissible spongiform encephalopathy was seen in several sections from each of a total of 1019 brains. We conclude that, for most practical purposes, Argentina may be considered to be free from BSE.