Atypical/Nor98 scrapie was first identified in 1998 in Norway. It is now considered as a worldwide disease of small ruminants and currently represents a significant part of the detected transmissible spongiform encephalopathies (TSE) cases in Europe. Atypical/Nor98 scrapie cases were reported in ARR/ARR sheep, which are highly resistant to BSE and other small ruminants TSE agents. The biology and pathogenesis of the Atypical/Nor98 scrapie agent in its natural host is still poorly understood. However, based on the absence of detectable abnormal PrP in peripheral tissues of affected individuals, human and animal exposure risk to this specific TSE agent has been considered low. In this study we demonstrate that infectivity can accumulate, even if no abnormal PrP is detectable, in lymphoid tissues, nerves, and muscles from natural and/or experimental Atypical/Nor98 scrapie cases. Evidence is provided that, in comparison to other TSE agents, samples containing Atypical/Nor98 scrapie infectivity could remain PrP^Sc negative. This feature will impact detection of Atypical/Nor98 scrapie cases in the field, and highlights the need to review current evaluations of the disease prevalence and potential transmissibility. Finally, an estimate is made of the infectivity loads accumulating in peripheral tissues in both Atypical/Nor98 and classical scrapie cases that currently enter the food chain. The results obtained indicate that dietary exposure risk to small ruminants TSE agents may be higher than commonly believed. Following the bovine spongiform encephalopathy (BSE) crisis and the identification of its zoonotic properties, a sanitary policy has been implemented based on both eradication of transmissible spongiform encephalopathies (TSE) in food-producing animals and exclusion of known infectious materials from the food chain. Atypical/Nor98 scrapie is a prion disease of small ruminants identified worldwide. Currently it represents a significant part of the TSE cases detected in Europe. The restricted tissue distribution of Atypical/Nor98 scrapie agent in its natural host and the low detected prevalence of secondary cases in affected flocks meant that it is believed to be a poorly transmissible disease. This has led to the view that Atypical/Nor98 scrapie is a spontaneous disorder for which human and animal exposure risk remains low. In this study we demonstrate that in affected individuals, Atypical/Nor98 scrapie agent can disseminate in lymphoid tissues, nerves, and muscles, challenging the idea that it is a brain-restricted infectious agent. Evidence for the deficiencies in the current methods applied for monitoring Atypical/Nor98 scrapie is provided that would indicate an underestimation in the prevalence in the general population and in the affected flocks. These elements challenge the hypothesis on the biology of this recently identified TSE agent.

Experimental transmission of bovine spongiform encephalopathy to sheep has prompted the implementation of a surveillance plan of scrapie in small ruminants by the European Union in all member states. Since its start over 30 000 animals have been tested, and the first seven cases of sheep with detectable PrP^res deposition in the central nervous system have been identified in Portugal. Notably, the pattern of PrP^res distribution in the brainstem was different from that previously described for scrapie and consistent in all seven animals. Moreover, the profile of the electrophoretic mobility of PrP^res after proteinase K treatment was equivalent in all cases analysed but distinct from that observed for scrapie. Notably, four animals had genotypes rarely associated with scrapie, including one animal homozygous for A₁₃₆R₁₅₄R₁₇₁. There were no cases found to exhibit vacuolation, a pattern of PrP^res distribution or PrP^res electrophoretic mobility corresponding to scrapie. These data reveal a putative atypical scrapie strain in Portugal not linked to specific Prnp genotypes.

Significance: The origin of transmissible BSE in cattle remains unestablished. Sheep scrapie is a potential source of this known zoonotic. Here we investigated the capacity of sheep scrapie to propagate in bovine PrP transgenic mice. Unexpectedly, transmission of atypical but not classical scrapie in bovine PrP mice resulted in propagation of classical BSE prions. Detection of prion seeding activity by in vitro protein misfolding cyclic amplification demonstrated BSE prions in the original atypical scrapie isolates. BSE prion seeding activity was also detected in ovine PrP mice inoculated with limiting dilutions of atypical scrapie. Our data demonstrate that classical BSE prions can emerge during intra- and interspecies passage of atypical scrapie and provide an unprecedented insight into the evolution of mammalian prions.

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 (PrP^C) into a misfolded pathological isoform (PrP^Sc or prion- proteinaceous infectious particle) that self-propagates by conformational conversion of PrP^C. Yet by an unknown mechanism, PrP^C can fold into different PrP^Sc 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 PrP^Sc 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).

Background: During the last decade, active surveillance for transmissible spongiform encephalopathies in small ruminants has been intensive in Europe. In many countries this has led to the detection of cases of atypical scrapie which, unlike classical scrapie, might not be contagious. EU legislation requires, that following detection of a scrapie case, control measures including further testing take place in affected flocks, including the culling of genotype susceptible to classical scrapie. This might result in the detection of additional cases. The aim of this study was to investigate the occurrence of additional cases in flocks affected by atypical scrapie using surveillance data collected in Europe in order to ascertain whether atypical scrapie, is contagious. Results: Questionnaires were used to collect, at national level, the results of active surveillance and testing associated with flock outbreaks in 12 European countries. The mean prevalence of atypical scrapie was 5.5 (5.0-6.0) cases per ten thousand in abattoir surveillance and 8.1 (7.3-9.0) cases per ten thousand in fallen stock. By using meta-analysis, on 11 out of the 12 countries, we found that the probability of detecting additional cases of atypical scrapie in positive flocks was similar to the probability observed in animals slaughtered for human consumption (odds ratio, OR = 1.07, CI_95%: 0.70-1.63) or among fallen stock (OR = 0.78, CI_95%: 0.51-1.2). In contrast, when comparing the two scrapie types, the probability of detecting additional cases in classical scrapie positive flocks was significantly higher than the probability of detecting additional cases in atypical scrapie positive flocks (OR = 32.4, CI_95%: 20.7-50.7). Conclusions: These results suggest that atypical scrapie is not contagious or has a very low transmissibility under natural conditions compared with classical scrapie. Furthermore this study stressed the importance of standardised data collection to make good use of the analyses undertaken by European countries in their efforts to control atypical and classical scrapie.

Active transmissible spongiform encephalopathy (TSE) surveillance in small ruminants across Europe was implemented in 2002 following the epizootic of bovine spongiform encephalopathy. Here, we report the potential emergence of classical scrapie in Portugal, in a background of enzootic atypical scrapie. Between 2003 and 2008, 375 459 small ruminants were screened in total, with 328 animals confirmed positive for NOR98 atypical scrapie. During this period, the prevalence rate of atypical scrapie for all years combined was 0.0874 % across the country. In this scenario, classical scrapie emerged as a single outbreak in 2008, with 12 identified cases. In contrast to other European countries, where classical scrapie has been enzootic for decades, these data indicate that, in Portugal, atypical scrapie is the predominant form of TSE. The findings reported here will have implications for the control of classical scrapie in Portugal, namely in terms of keeping the country free of enzootic classical scrapie.