Bovine Spongiform Encephalopathy Infected Cattle Research Articles

Use of bovine recombinant prion protein and real-time quaking-induced conversion to detect cattle transmissible mink encephalopathy prions and discriminate classical and atypical L- and H-Type bovine spongiform encephalopathy.

Prions are amyloid-forming proteins that cause transmissible spongiform encephalopathies through a process involving conversion from the normal cellular prion protein to the pathogenic misfolded conformation (PrPSc). This conversion has been used for in vitro assays including serial protein misfolding amplification and real-time quaking induced conversion (RT-QuIC). RT-QuIC can be used for the detection of prions in a variety of biological tissues from humans and animals. Extensive work has been done to demonstrate that RT-QuIC is a rapid, specific, and highly sensitive prion detection assay. RT-QuIC uses recombinant prion protein to detect minute amounts of PrPSc. RT-QuIC has been successfully used to detect PrPSc from different prion diseases with a variety of substrates including hamster, human, sheep, bank vole, bovine and chimeric forms of prion protein. However, recombinant bovine prion protein has not been used to detect transmissible mink encephalopathy (TME) or to differentiate types of bovine spongiform encephalopathy (BSE) in samples from cattle. We evaluated whether PrPSc from TME and BSE infected cattle can be detected with RT-QuIC using recombinant bovine prion proteins, and optimized the reaction conditions to specifically detect cattle TME and to discriminate between classical and atypical BSE by conversion efficiency. We also found that substrate composed of the disease associated E211K mutant protein can be effective for the detection of TME in cattle and that wild type prion protein appears to be a practical substrate to discriminate between the different types of BSEs.

Scientific Opinion on BSE risk in bovine intestines and mesentery

Bovine intestines and mesenteries in the European Union (EU) have to be removed from the food and feed chain. The opinion provides a quantitative assessment of the Bovine Spongiform Encephalopathy (BSE) infectious load that might enter the food and feed chain yearly if bovine intestine and mesentery from animals born and raised in the EU would be re-allowed for consumption. Data on the evolution of the BSE infectious titre; and of the weight of histological structures accumulating BSE infectivity, were collected. The Cattle TSE Monitoring Model (C-TSEMM) was used to estimate the number of BSE infected cattle entering undetected in the food and feed chain yearly. A model named TSEi was developed to estimates the BSE infectious load in tissues from infected animals at different ages and the total yearly infectious load that could enter the food and feed chain in the EU27. In BSE infected cattle, the infectivity associated with intestine and mesentery reaches its maximum in animals younger than 18 months and then progressively declines to a minimum value in animals older than 60 months. Due to the decline of the BSE prevalence in the EU, between 2007 and 2012, the yearly amount of BSE infectivity associated with intestine and mesentery (sent to destruction) from animals entering the food and feed chain was reduced by a factor of 10. However, over this period, the maximum level of exposure to the BSE agent for individuals that would have consumed these tissues remained stable. Finally, the TSEi model indicated that the removal of the last four metres of the small intestine and of the caecum from the food and feed chain would result in a major reduction of the BSE exposure risk associated with intestine and mesentery in cattle.

Complementary studies detecting classical bovine spongiform encephalopathy infectivity in jejunum, ileum and ileocaecal junction in incubating cattle

Recently we have described the distribution of bovine spongiform encephalopathy (BSE) infectivity and/or PrPSc in Peyer’s patches (PP) of the small intestine of orally BSE infected cattle. In this follow-up study additional jejunal and ileal PP’s and ileocaecal-junction tissue samples from 1, 4, and 24 months post infection (mpi) were examined by mouse (Tgbov XV) bioassay. Infectivity was demonstrated in ileal PP’s 4 mpi and the distribution/extent of infectivity at 24 mpi was comparable to those seen at earlier time points, revealing no indication for a decline/clearance. These data are relevant for the definition of Specified Risk Materials in the context of the TSE legislation worldwide.

Prion in Saliva of Bovine Spongiform Encephalopathy–Infected Cattle

Prion in Saliva of Bovine Spongiform Encephalopathy–Infected Cattle

Prion in Saliva of Bovine Spongiform Encephalopathy–Infected Cattle

Prion in Saliva of Bovine Spongiform Encephalopathy–Infected Cattle

Minimal Involvement of the Circumventricular Organs in the Pathogenesis of Spontaneously Arising and Experimentally Induced Classical Bovine Spongiform Encephalopathy

In sheep infected experimentally with the bovine spongiform encephalopathy (BSE) agent, amplification of infectivity in peripheral organs during early preclinical stages is thought to contribute to high titres of the agent being detected in blood, with subsequent haematogenous neuroinvasion through the circumventricular organs (CVOs). In contrast, little disease-associated prion protein (PrP(d)) or infectivity is detected in the peripheral tissues of cattle during the preclinical and clinical stages of BSE. The aim of this study was to investigate immunohistochemically the role of haematogenous neuroinvasion in cattle with spontaneously arising and experimentally induced BSE. There was almost complete absence of PrP(d) in the peripheral organs of BSE infected cattle. Additionally, there was minimal involvement of the CVOs during preclinical disease and there was progressive caudorostral accumulation of PrP(d) in the brain. These findings do not support haematogenous neuroinvasion in the bovine disease.

BSE infectivity in jejunum, ileum and ileocaecal junction of incubating cattle

To establish bovine spongiform encephalopathy (BSE) public health protection measures it is important to precisely define the cattle tissues considered as specified risk materials (SRM). To date, in pre-clinical BSE infected cattle, no evidence of the BSE agent had been found in the gut outside of the ileal Peyer's Patches. This study was undertaken to determine when and where the pathological prion protein (PrPSc) and/or BSE infectivity can be found in the small intestine of cattle 4 to 6 months of age, orally challenged with BSE. Samples of the jejunum, the ileum and the ileocaecal junction from 46 BSE infected cattle, culled from 1 up to 44 months post infection (mpi) were examined by immunohistochemistry. Samples from cattle 8 mpi to 20 mpi were additionally studied by PTA Western blot, rapid tests, and by mouse (TgbovXV) bioassay. In doing so nearly all of the cattle, from 4 up to 44 mpi, had detectable amounts of PrPSc and/or infectivity in the distal ileum. In the distal ileum clear time-dependent variations were visible concerning the amount of PrPSc, the tissue structures affected, and the cells involved. BSE infectivity was found not only in the ileum and ileocaecal junction but also in the jejunum. The systematic approach of this study provides new data for qualitative and quantitative risk assessments and allows defining bovine SRM more precisely.

Scientific Opinion on the revision of the quantitative risk assessment (QRA) of the BSE risk posed by processed animal proteins (PAPs)

The cattle Bovine Spongiform Encephalopathy (BSE) risk posed by bovine derived Processed Animal Proteins (PAPs) in feed was estimated, the diagnostic methods and their sensitivity to detect animal proteins in feed and compared different risk assessment methods for animal proteins in feed was reviewed. It was concluded that the current global limit of detection for PAPs in feed is still considered to be 0.1%. Data related to BSE monitoring and PAP production in the European Union (EU) were considered. A model (EFSA QRA PAP model) was developed to study the magnitude of the total BSE infectivity in PAPs and in ruminant feed under a certain scenario assuming some specific cross-contaminations. On the basis of the 2009 BSE surveillance data in the EU, assuming a 0.1% contamination (the limit of detection for PAP in feed) with non-ruminant PAPs and according to the EFSA QRA PAP model, the total BSE infectivity load that could enter in cattle feed per year in the EU would be equivalent to 0.2 Cattle oral Infectious Dose 50% (Co ID50) (9 x 10-5 - 1.3 CI95%) (that would mean that less than one additional BSE infected cattle could be expected in the EU cattle population per year with an upper 95% confidence). The specific scenario described by the model and the related assumptions and uncertainties are discussed in this scientific opinion.

A single step multiplex immunofluorometric assay for differential diagnosis of BSE and scrapie

Although there is no evidence that the European sheep population has been infected with bovine spongiform encephalopathy (BSE), distinguishing this from scrapie is paramount, given the association between BSE exposure and the human transmissible spongiform encephalopathy (TSE), variant Creutzfeldt-Jakob disease. The capability to differentially diagnose TSEs in sheep is thus essential in order to safeguard the food chain and human health. Biochemical methods for differentiating BSE and scrapie are largely reliant on assessment by Western blot (WB) analysis of the abnormal disease associated prion protein PrP(D) following partial proteolytic digestion. WB banding patterns obtained using a panel of antibodies enable different strain specific conformations of PrP(D) to be distinguished. This approach provides a robust confirmatory test but one which is not appropriate for high throughput screening. A simple, one step, bead array flow cytometry based multiplex immunofluorometric assay has been developed which is suitable for simultaneous screening and confirmation. Using a combination of antibodies directed towards three PrP epitopes enabled differential diagnosis of scrapie and BSE. Proof of principle studies indicated a high predictive value (100%) when applied to brain samples from control animals, BSE infected cattle and sheep naturally infected with scrapie or experimentally infected with BSE.

The identification of disease-induced biomarkers in the urine of BSE infected cattle

BackgroundThe bovine spongiform encephalopathy (BSE) epidemic and the emergence of a new human variant of Creutzfeldt-Jakob Disease (vCJD) have led to profound changes in the production and trade of agricultural goods. The rapid tests currently approved for BSE monitoring in slaughtered cattle are all based on the detection of the disease related isoform of the prion protein, PrPd, in brain tissue and consequently are only suitable for post-mortem diagnosis. Objectives: In instances such as assessing the health of breeding stock for export purposes where post-mortem testing is not an option, there is a demand for an ante-mortem test based on a matrix or body fluid that would permit easy access and repeated sampling. Urine and urine based analyses would meet these requirements.ResultsTwo dimensional differential gel eletrophoresis (2D-DIGE) and mass spectrometry analyses were used to identify proteins exhibiting differential abundance in the urine of BSE infected cattle and age matched controls over the course of the disease. Multivariate analyses of protein expression data identified a single protein able to discriminate, with 100% accuracy, control from infected samples. In addition, a subset of proteins were able to predict with 85% ± 13.2 accuracy the time post infection that the samples were collected.ConclusionThese results suggest that in principle it is possible to identify biomarkers in urine useful in the diagnosis, prognosis and monitoring of disease progression of transmissible spongiform encephalopathy diseases (TSEs).

Stress response in the central nervous system of a transgenic mouse model of bovine spongiform encephalopathy

An immunohistochemical study was performed to evaluate the stress-related proteins heat shock protein 25 (HSP25) and metallothionein 1 + 2 (MT1 + 2) in the brains of a murine model of bovine spongiform encephalopathy (BSE). Transgenic mice (BoTg110) expressing the bovine cellular prion protein were intracerebrally inoculated with brainstem homogenate from BSE infected cattle. PrP BSE deposits were found in the brain as early as 150 days post-inoculation (dpi) and in mice sacrificed terminally at 290–320 dpi. Glial proliferation and spongiform change were associated with an increase in glial immunostaining of MT1 + 2 and HSP25, respectively. These proteins are associated with oxidative stress and heavy metal metabolism, which may have a role in the pathogenesis of BSE.

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.

Application of systems analysis in modelling the risk of bovine spongiform encephalopathy (BSE)

Bovine spongiform encephalopathy (BSE), widely known as “mad cow disease”, has virtually crippled the British livestock industry. Even though, no cases of BSE have been reported in the United States (US), a similar epidemic in the US would be catastrophic. The added concern for the risk of introduction of the human disease called variant Creutzfeldt‐Jacob disease that has been linked to eating meat of BSE infected cattle compounds the risk of BSE. Systems dynamics models based on the underlying transmission pathways of BSE can help to anticipate the spread of this disease in different cattle populations and assist in the evaluation of potential risk mitigations for preventing its introduction or controlling its spread if it was introduced. With this in mind, an age and health status structured systems dynamics model was developed. By making assumptions and setting up feasible scenarios, the model can be used to examine potential prevalence and incidences rates of BSE; the effect of mitigations including changes in feeding habits or rendering processes and/or other policies and regulations designed to prevent the introduction of BSE. The systems dynamics simulation model enabled us to create virtual experiments whose real‐world analogues would otherwise be expensive, dangerous, or even impossible to carryout.