Molecular Strain Typing and Environmental Persistence of Ruminant Transmissible Spongiform Encephalopathies

A novel thermolysin digestion method for the molecular strain-typing of ruminant TSEs has been developed which resulted in the clearance of cellular prion protein (PrP) from healthy sheep or cattle brain homogenates, while digestion of scrapie or BSE infected samples resulted in the generation of the full-length disease-related isoform, PrP. Using antibodies against the amino terminal region of PrP it was possible to distinguish ovine scrapie from ovine BSE, permitting the potential identification of BSE infected sheep within the UK flock. The identification of a disease-associated, endogenously-generated fragment of PrP (termed C2) in scrapie infected sheep is also described. Absent in healthy brain homogenates, the neuroanatomical distribution of both C2 fragments and thermolysin-resistant PrP permitted the classification of four groups within a sample of natural scrapie cases which may be representative of scrapie strain heterogeneity in the UK. The retention of ovine scrapie and bovine BSE prion protein during incubation with six UK lowland soils in soil-packed columns over a period of 18 months was also monitored. Data was collected on the persistence, the vertical migration, and the distribution of PrP in soil component fractions. PrP bound to all six soils, with elution being dependent on soil type. The majority of PrP bound irreversibly, and no migration of PrP was observed. Differences in PrP persistence were dependent on soil type and prion strain. PrP persistence on a single soil at defined pH, temperature, or moisture contents indicated the initial deposition of PrP within the column was dependent on soil pH and persistence was inversely correlated to temperature. Reduced soil moisture content resulted in increased elution and persistence of PrP. Data suggests that the interaction of PrP with soil is a complex phenomenon dependent not only upon soil type and TSE strain but also environmental factors such as soil temperature, pH, and moisture content.