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In view of the established link between bovine spongiform encephalopathy (BSE) and variant Creutzfeldt–Jakob disease and of the susceptibility of sheep to experimental BSE, the detection of potential cases of naturally occurring BSE in sheep has become of great importance. In this study, the immunohistochemical (IHC) phenotype of disease-associated prion protein (PrPd) accumulation has been determined in the brain of 64 sheep, of various breeds and PrP genotypes, that had developed neurological disease after experimental BSE challenge with different inocula by a range of routes. Sheep BSE was characterized by neuron-associated intra- and extracellular PrPd aggregates and by conspicuous and consistent deposits in the cytoplasm of microglia-like cells. The stellate PrPd type was also prominent in most brain areas and marked linear deposits in the striatum and midbrain were distinctive. Sheep of the ARR/ARR and ARQ/AHQ genotypes displayed lower levels of PrPd than other sheep, and intracerebral BSE challenge resulted in higher levels of PrPd accumulating in the brain compared with other routes. The PrP genotype and the route of challenge also appeared to affect the incubation period of the disease, giving rise to complex combinations of magnitude of PrPd accumulation and incubation period. Despite these differences, the phenotype of PrPd accumulation was found to be very consistent across the different factors tested (notably after subpassage of BSE in sheep), thus highlighting the importance of detailed IHC examination of the brain of clinically affected sheep for the identification of potential naturally occurring ovine BSE.

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BackgroundThe risk of scrapie infection increases with increased duration and proximity of contact between sheep at lambing. Scrapie infectivity has not been detected in milk but cellular prion protein, the precursor of disease-associated prion protein PrPd, has been found in milk from ruminants. To determine whether milk is able to transmit scrapie, 18 lambs with a prion protein genotype associated with high susceptibility to scrapie (VRQ/VRQ) were fed milk from twelve scrapie-affected ewes of the same genotype, and 15 VRQ/VRQ sheep reared on scrapie-free dams served as controls.ResultsThree lambs fed milk from scrapie-affected ewes were culled due to intercurrent diseases at 43, 44 and 105 days of age respectively, and PrPd was detected in the distal ileum of the first two lambs, whilst PrPd was not found in lymphoreticular tissues in the third lamb. A control lamb, housed in a separate pen and culled at 38 days of age, was also negative for PrPd in a range of tissues. Samples of recto-anal mucosa associated lymphoid tissue collected from the remaining 15 live lambs at seven months of age (between five to seven months after mixing) were positive for PrPd in the scrapie milk recipients, whereas PrPd was not detected in the remaining 14 controls at that time. A subsequent sample collected from control lambs revealed PrPd accumulation in two of five lambs eight months after mixing with scrapie milk recipients suggestive of an early stage of infection via lateral transmission. By contrast, the control sheep housed in the same building but not mixed with the scrapie milk recipients were still negative for PrPd.ConclusionThe presence of PrPd in distal ileum and rectal mucosa indicates transmission of scrapie from ewe to lamb via milk (or colostrum) although it is not yet clear if such cases would go on to develop clinical disease. The high level of infection in scrapie-milk recipients revealed by rectal mucosal testing at approximately seven months of age may be enhanced or supplemented by intra-recipient infection as these lambs were mixed together after feeding with milk from scrapie-affected ewes and we also observed lateral transmission from these animals to lambs weaned from scrapie-free ewes.

Sixty Romney sheep of three prion protein genotypes were dosed orally at six months of age with an inoculum prepared from the brains of cattle clinically affected with BSE, and 15 sheep were left undosed as controls. They were randomly assigned within genotype to groups and were sequentially euthanased and examined postmortem at intervals of six or 12 months, depending on their predicted susceptibility. Tissue pools prepared from the three, four or five dosed animals in each group were inoculated into groups of 20 RIII mice as a bioassay for infectivity. Separate inocula were prepared from the matched control sheep killed at each time. In the ARQ/ARQ sheep killed four months after inoculation, infectivity was detected in the Peyer's patch tissue pool, and at 10 months it was detected in the spleen pool; from 16 months, infectivity was detected in a range of nervous and lymphoreticular tissues, including the spinal cord pool, distal ileum excluding Peyer's patches, liver, Peyer's patches, mesenteric and prescapular lymph nodes, spleen, tonsil and cervical thymus. No infectivity was detected in the tissue pools from the ARQ/ARR and ARR/ARR sheep killed 10 months or 22 months after infection.