BSE in Great Britain: consistency of the neurohistopathological findings in two random annual samples of clinically suspect cases

In several EU member states, bovine spongiform encephalopathy (BSE) cases have been identified in cattle born after the reinforced ban (BARB cases), for reasons that are not entirely clear. Epidemiological...

BOVINE spongiform encephalopathy (BSE) is a transmissible spongiform encephalopathy (TSE) that during the last three decades has overturned all the paradigms of surveillance, diagnosis, legislation and public concern. Several origins...

For almost two decades after the discovery of the first bovine spongiform encephalopathy (BSE) case, it was generally accepted that only one BSE strain existed globally. However, in 2004, two novel BSE forms (L-type and H-type) were separately identified in two different European Member States, forms that differed from the classical (C-type) form by their biochemical properties and by the pattern of PrPSc deposition as determined by immunohistochemistry (IHC). 60 atypical BSE cases have been identified worldwide as of November 2010, including one H- and one L-type BSE case each in Germany. However, it was not known whether the biological properties (pathogenesis and agent distribution, as well as transmissibility to other species) of these novel forms were the same as in classical BSE cases. Eleven calves were thus challenged intracranially, five with the German H-type and six with German L-type BSE cases. The experimental design and the clinical studies, followed by laboratory testing, are described in this manuscript.

Bovine spongiform encephalopathy (BSE) was first identified in Great Britain (GB) in 1986 and was subsequently detected in many other countries, worldwide. A decade after the start of the bovine epidemic, the first cases of new variant Creutzfeldt-Jakob disease (vCJD) in humans were linked to probable ingestion of BSE infected tissue, highlighting a new zoonotic disease. An abnormal protease-resistant protein (PrP(res)) in a diseased subject, derived from a post-translational change of a normal host cellular membrane protein (PrP(c)), is a reliable disease marker for the whole group of neurodegenerative transmissible spongiform encephalopathies (TSEs). Immunology-based techniques, such as Western immunoblotting, have previously indicated that BSE cases all give a uniform molecular profile for PrP(res). Periodic lesion profiling of the spongiform change throughout different brain regions of infected mice and cattle has also indicated a single agent for BSE. However, in 2001 rapid testing for PrP(res) was introduced for the active surveillance of ruminants within Europe, and approximately 40 BSE cases have now been recognized that differ in their molecular profiles from those typically found. These unusual BSE cases have been detected in several European countries, and in Japan and the USA. At present, the cases appear as two distinct types based on the molecular mass (Mm) of the unglycosylated PrP(res) protein band relative to that of classical BSE. One type is of a higher Mm (H-type) and the other shows a lower Mm (L-type). Transmission studies in mice have shown that both H-type and L-type BSE have biological characteristics that are different from those of the classical BSE agent. This study describes the prion protein (PRNP) genotype and molecular profiles of the first two cases of H-type BSE detected in GB in comparison with those obtained for classical BSE, scrapie in sheep from GB and a control H-type BSE case from France.

Bovine spongiform encephalopathy

Cross-contamination of cattle feed with meat and bone meal (MBM) allowed in feed for other species is regarded as the current hypothesis for the infection pathway of Bovine Spongiform Encephalopathy (BSE) cases occurring after the implementation of a ban on feeding MBM to cattle. This study was aimed at establishing a spatial relation between BSE cases in Switzerland and the findings of MBM in cattle feed. A cluster analysis and a cohort study were performed. Two hundred sixteen BSE cases born after December 1990 and detected until August 1st 2005, screening data of 504 feed producers between 1996 and 2001 and population data from the Swiss 2001 cattle census were included. The cluster analysis showed feed producer, positive for MBM contaminations in cattle feed, as possible cluster centres for BSE cases. In the cohort study, farms within a radius of 2 and 10 km around positive feed producers showed significantly higher odds to have a BSE case than the control group. The odds ratio and its 95% confidence interval were 2.23 (1.26-3.93) for the 2 km radius and 1.38 (1-1.9) for the 10 km radius. The results provide evidence for a spatial relation between cross-contamination and BSE occurrence. These findings support the hypothesis of cross-contamination to be an important route for BSE transmission after a feed ban.

Interest in the proper neuropathological and molecular characterization of bovine spongiform encephalopathy (BSE) has increased since asymptomatic and atypical cases were detected in the cattle population by active disease surveillance. In this respect we investigated a total of 95 confirmed BSE cases originating from different active and passive surveillance categories (clinical suspects, emergency-slaughter, fallen stock and routinely slaughter) in Switzerland for their neuropathological and molecular phenotype. We looked for measurable differences between these categories in lesion profile, severity of spongiform change, degree of astrocytosis as well as immunohistochemical and molecular patterns of the disease-associated isoform of the prion protein (PrPd) in the caudal brainstem. Our results indicate significantly higher intensities of spongiform change in clinically affected compared to asymptomatic BSE cases. Similar effects were in trend observed for the intensities of PrPd deposition and astrocytosis, whereas the frequencies of morphological PrPd types and the molecular patterns in Western immunoblot were not different. Importantly, none of the animals included in this study revealed features of atypical BSE. Taken together, this study suggests that both clinically affected as well as asymptomatic Swiss BSE cases in cattle share the neuropathological and molecular phenotype of classical BSE and that asymptomatic classical BSE cases are at a pre-clinical stage of the disease rather than representing a true sub-clinical form of BSE.

The epidemiology and possibly the etiology of bovine spongiform encephalopathy (BSE) have recently been recognized to be heterogeneous. In particular, three types [classical (C) and two atypical (H, L)] have been identified, largely on the basis of characteristics of the proteinase K (PK)-resistant core of the misfolded prion protein associated with the disease (PrPres). The present study was conducted to characterize the 17 Canadian BSE cases which occurred prior to November 2009 based on the molecular and biochemical properties of their PrPres, including immunoreactivity, molecular weight, glycoform profile and relative PK sensitivity. Two cases exhibited molecular weight and glycoform profiles similar to those of previously reported atypical cases, one corresponding to H-type BSE (case 6) and the other to L-type BSE (case 11). All other cases were classified as C-type. PK digestion under mild and stringent conditions revealed a reduced protease resistance in both of these cases compared to the C-type cases. With Western immunoblotting, N-terminal-specific antibodies bound to PrPres from case 6 but not to that from case 11 or C-type cases. C-terminal-specific antibodies revealed a shift in the glycoform profile and detected a fourth protein fragment in case 6, indicative of two PrPres subpopulations in H-type BSE. No mutations suggesting a genetic etiology were found in any of the 17 animals by sequencing the full PrP-coding sequence in exon 3 of the PRNP gene. Thus, each of the three known BSE types have been confirmed in Canadian cattle and show molecular characteristics highly similar to those of classical and atypical BSE cases described from Europe, Japan and the USA. The occurrence of atypical cases of BSE in countries such as Canada with low BSE prevalence and transmission risk argues for the occurrence of sporadic forms of BSE worldwide.

Targeted screening of high-risk cattle populations for BSE to augment mandatory reporting of clinical suspects

Science, policy, and politics: the case of BSE

Evaluation of a Rapid Western Immunoblotting Procedure for the Diagnosis of Bovine Spongiform Encephalopathy (BSE) in the UK

Variant Creutzfeldt-Jakob disease (vCJD) has been recognized to date only in individuals homozygous for methionine at PRNP codon 129. Here we show that transgenic mice expressing human PrP methionine 129, inoculated with either bovine spongiform encephalopathy (BSE) or variant CJD prions, may develop the neuropathological and molecular phenotype of vCJD, consistent with these diseases being caused by the same prion strain. Surprisingly, however, BSE transmission to these transgenic mice, in addition to producing a vCJD-like phenotype, can also result in a distinct molecular phenotype that is indistinguishable from that of sporadic CJD with PrP(Sc) type 2. These data suggest that more than one BSE-derived prion strain might infect humans; it is therefore possible that some patients with a phenotype consistent with sporadic CJD may have a disease arising from BSE exposure.

After the discovery of variant Creutzfeldt-Jakob disease (vCJD), scientific advances quickly led to post-mortem tests to identify late-stage bovine spongiform encephalopathy (BSE) disease. These were first used in Switzerland in 1999 for active BSE surveillance of a) fallen and emergency-slaughter bovines (risk stock) and b) 5% sample of routinely slaughtered cattle over 30 months of age. In 1999 and 2000, Switzerland's estimated 103 BSE positives per 1000000 adult cattle put it in the same BSE risk classification as UK and Portugal. In July 2000, the European Union's Scientific Steering Committee published its methodology (and first vetted results) for geographical BSE risk (GBR) assessment in cattle. Member states with no BSE cases found themselves, on rational assessment, classified as GBR III (BSE likely but not confirmed, or confirmed at a lower level). Because of Europe's thus highly assessed BSE risks, active BSE surveillance of adult cattle in all member states began in January 2001 using one of three validated post-mortem tests. Implementation was variable across member states in January to March 2001 but, where operational, active surveillance was typically achieved for around 13300 routinely slaughtered and 1000 risk stock per month per 1000000 adult cattle; BSE positive rates were 60 and 600 per 1000000 routinely slaughtered and risk cattle, respectively. By the second half of 2001, active BSE surveillance was operating reasonably in most member states, although anomalies persisted. Performance and results for July to December 2001 and for January to June 2002 are considered in detail. The BSE positive rate decreased substantially in UK, Portugal and Ireland between semesters, whereas Spain's rates increased for both routinely slaughtered and risk bovines. Based on 1450000 routinely slaughtered and 135000 risk stock as standard, France could have expected 153 BSE positives in July to December 2001 (109 in January to June 2002); Italy 154 (67); and Germany only 39 (48). When sample-based surveillance data were scaled up and combined with clinical BSE cases, Great Britain's BSE positives were estimated at around 400 per 1000000 adult cattle in 2002 compared with over 1000 per 1000000 adult cattle in 2000. Age distributions for cattle subject to active BSE surveillance have been underexploited. The major transmissible spongiform encephalopathy (TSE) which affects sheep and goats is scrapie. Passive surveillance of scrapie is associated with substantial under-reporting. Susceptibility to scrapie depends strongly on sheep genotype; but resistance to scrapie does not necessarily confer resistance of sheep to BSE. Because of uncertainty about the true prevalence of scrapie-infected adult sheep and concern that BSE in sheep may be missed, the European Union pre-empted its planned evaluation of rapid post-mortem TSE tests in sheep by requiring the rapid TSE testing of small ruminants from April 2002 with one of the three cattle-validated tests. Basic requirements for active TSE surveillance in sheep were: random sample of 6000 fallen sheep and of 60000 routinely slaughtered adult native sheep to be tested per member state by end March 2003. Lower surveillance targets were set for countries with under 1000000 adult sheep. Adequately to map scrapie-susceptible genotypes and identify resistant genotypes, a random sample of 500 routinely slaughtered native adult sheep was to be genotyped, together with each TSE rapid test positive adult sheep and two sets of three suitably sampled controls. By the end of August 2002, when 41% of the initial surveillance time had elapsed, only 20% of the European joint target for routinely slaughtered adult sheep had been completed, but that for fallen sheep was exceeded. Except in Ireland, the upper 95% confidence bound on TSE prevalence exceeded 500 per 1000000 routinely slaughtered adult sheep in reporting-compliant countries with more than 1000000 adult sheep. The UK, Greece, Italy and France were likely to approach the goal of 100 TSE rapid test positives on completion of their assigned first-year surveillance target for sheep. Results from the recommended genotyping of TSE positive adult sheep and controls for use in inferring differential TSE-positive susceptibility by genotype are awaited. Only by genotyping 5000-50000 TSE-positive adult sheep, a massive undertaking even on the European scale, will it become clear whether scrapie resistance is relative rather than absolute. This paper details Europe's quantitative evolution in TSE surveillance.

Variant Creutzfeldt-Jakob disease: French versus British.

Ten years of BSE surveillance in Italy: Neuropathological findings in clinically suspected cases