PrPSc spreading patterns in the brain of sheep linked to different prion types

Scrapie in sheep and goats has been known for more than 250 years and belongs nowadays to the so-called prion diseases that also include e.g. bovine spongiform encephalopathy in cattle (BSE) and Creutzfeldt-Jakob disease in humans. According to the prion hypothesis, the pathological isoform (PrPSc) of the cellular prion protein (PrPc) comprises the essential, if not exclusive, component of the transmissible agent. Currently, two types of scrapie disease are known - classical and atypical/Nor98 scrapie. In the present study we examine 24 cases of classical and 25 cases of atypical/Nor98 scrapie with the sensitive PET blot method and validate the results with conventional immunohistochemistry. The sequential detection of PrPSc aggregates in the CNS of classical scrapie sheep implies that after neuroinvasion a spread from spinal cord and obex to the cerebellum, diencephalon and frontal cortex via the rostral brainstem takes place. We categorize the spread of PrPSc into four stages: the CNS entry stage, the brainstem stage, the cruciate sulcus stage and finally the basal ganglia stage. Such a sequential development of PrPSc was not detectable upon analysis of the present atypical/Nor98 scrapie cases. PrPSc distribution in one case of atypical/Nor98 scrapie in a presumably early disease phase suggests that the spread of PrPSc aggregates starts in the di- or telencephalon. In addition to the spontaneous generation of PrPSc, an uptake of the infectious agent into the brain, that bypasses the brainstem and starts its accumulation in the thalamus, needs to be taken into consideration for atypical/Nor98 scrapie

Unexpected high frequency of neurofibroma in the celiac ganglion of German cattle

In a study originally designed to find potential risk factors for bovine spongiform encephalopathy (BSE) we examined tissues from 403 Holstein Frisian cattle in total. These included 20 BSE cattle and their 236 birth- and feeding cohort animals plus 32 offspring, 103 age, breed and district-matched control cattle and further twelve cattle with neurological signs. In addition to the obex, we examined the celiac ganglion, cervical cranial ganglion, trigeminal ganglion and proximal ganglion of the vagus nerve using histological techniques. Unexpectedly, we found a high number of neurofibroma, a benign peripheral nerve sheath tumor consisting of Schwann cells, fibroblasts and perineural cells. The neurofibroma were present only in the celiac ganglion and found during histologic examination. With a frequency of 9.91% in BSE cattle and their cohorts (case animals) and 9.09% in the age, breed and district matched control animals there seems to be no correlation between the occurrence of BSE and neurofibroma. Benign peripheral nerve sheath tumors have been described more often in cattle than in other domestic animals. Usually, they are incidental macroscopic findings in the thoracic ganglia during meat inspection. To our knowledge, there are no previous systematic histologic studies including bovine celiac ganglia at all. The high incidence of celiac ganglia neurofibroma may play a role in the frequently occurring abomasal displacements in Holstein Frisian cattle as the tumors might cause a gastrointestinal motility disorder. At present a genetic predisposition for these neoplasms cannot be ruled out.Grants ZN 1294 and ZN 2168, Volkswagen Stiftung financed the first years of sample collection, i.e. material from BSE and cohort animals regarding staff and material expenses Project number: 38028266, Deutsche Forschungsgemeinschaft (DFG), was granted for the original study design to identify risk factors for BSE in the cohort animals of BSE animalsSaarland University within the funding programme Open Access Publishing

Accumulation of Pathological Prion Protein PrPSc in the Skin of Animals with Experimental and Natural Scrapie

Prion infectivity and its molecular marker, the pathological prion protein PrPSc, accumulate in the central nervous system and often also in lymphoid tissue of animals or humans affected by transmissible spongiform encephalopathies. Recently, PrPSc was found in tissues previously considered not to be invaded by prions (e.g., skeletal muscles). Here, we address the question of whether prions target the skin and show widespread PrPSc deposition in this organ in hamsters perorally or parenterally challenged with scrapie. In hamsters fed with scrapie, PrPSc was detected before the onset of symptoms, but the bulk of skin-associated PrPSc accumulated in the clinical phase. PrPSc was localized in nerve fibres within the skin but not in keratinocytes, and the deposition of PrPSc in skin showed no dependence from the route of infection and lymphotropic dissemination. The data indicated a neurally mediated centrifugal spread of prions to the skin. Furthermore, in a follow-up study, we examined sheep naturally infected with scrapie and detected PrPSc by Western blotting in skin samples from two out of five animals. Our findings point to the skin as a potential reservoir of prions, which should be further investigated in relation to disease transmission

Accuracy of Subclinical Ketosis Detection with Rapid Test Methods for BHBA in Blood in Commercial Dairy Farms

This study gives an overview of the performance and accuracy of devices used for the fast measurement of β-hydroxybutyrate (BHBA) in blood for the on-farm indication of subclinical ketosis. Data were collected on ten dairy farms. In each farm, blood samples were taken from ten cows on four test days (2, 4, 9 and 11), resulting in 400 samples. The reference method was the BHBA concentration in blood serum (BHBALAB). Four different devices that measure BHBA in whole blood were tested. The thresholds applied for identifying subclinical ketosis were ≥1.0, ≥1.2 and ≥1.4 mmol/L in blood serum. The BHBALAB was assigned in three classes: low—≤0.9 mmol/L; high—>0.9 mmol/L; and total—all values unclassified. Due to initial negative effects on the health and performance of cows with BHBA levels ≥0.9 mmol/L, this cut-off was chosen. The Passing–Bablok regression revealed different constant as well as absolute biases for each device in the aforementioned classes. The area under the receiver operating characteristics curve indicated highly accurate results, with 94–97% accuracy levels. As an overall conclusion, the performance of the devices was good and supports their use by farmers for the detection of subclinical ketotic cows in their herds

Bov-tA Short Interspersed Nucleotide Element Sequences in Circulating Nucleic Acids from Sera of Cattle with Bovine Spongiform Encephalopathy (BSE) and Sera of Cattle Exposed to BSE

Circulating nucleic acids (CNA) are known to be enriched in repetitive DNA sequences in humans. Here, bovine sera CNA were analyzed to determine if cell stress-related short interspersed nucleotide elements (SINEs) could be detected in sera from cattle associated with bovine spongiform encephalopathy (BSE). Nucleic acids were extracted, amplified, cloned, and sequenced from the sera of protease-resistant prion protein (PrP(res))-positive cattle (n = 2) and sera from BSE-cohort cows (n = 6); 150 out of 163 clones revealed the presence of, on average, an 80-bp sequence from the 3′ region of Bov-tA SINE. A PCR protocol was developed that differentially identified SINE-associated CNA in BSE-exposed versus normal cattle. CNA were extracted from a serum vesicular fraction after controlled blood collection and processing procedures. Sera from four confirmed cases of BSE, 137 BSE-exposed cohort animals associated with eight confirmed BSE cases, and 845 healthy, PrP(res)-negative control cows were tested. All four sera from confirmed BSE cases were repeatedly reactive in the assay. BSE-exposed cohorts had a 100-fold higher occurrence of repeatedly reactive individuals per cohort (average = 63%; range = 33% to 91%), compared to healthy controls (average = 0.6%; P < 0.001). This study shows that BSE-confirmed and cohort animals possess a unique profile of SINE-associated serum CNA that can be utilized as a marker that highly correlates to BSE exposure

Accuracy of Subclinical Ketosis Detection with Rapid Test Methods for BHBA in Blood in Commercial Dairy Farms

This study gives an overview of the performance and accuracy of devices used for the fast measurement of β-hydroxybutyrate (BHBA) in blood for the on-farm indication of subclinical ketosis. Data were collected on ten dairy farms. In each farm, blood samples were taken from ten cows on four test days (2, 4, 9 and 11), resulting in 400 samples. The reference method was the BHBA concentration in blood serum (BHBALAB). Four different devices that measure BHBA in whole blood were tested. The thresholds applied for identifying subclinical ketosis were ≥1.0, ≥1.2 and ≥1.4 mmol/L in blood serum. The BHBALAB was assigned in three classes: low—≤0.9 mmol/L; high—>0.9 mmol/L; and total—all values unclassified. Due to initial negative effects on the health and performance of cows with BHBA levels ≥0.9 mmol/L, this cut-off was chosen. The Passing–Bablok regression revealed different constant as well as absolute biases for each device in the aforementioned classes. The area under the receiver operating characteristics curve indicated highly accurate results, with 94–97% accuracy levels. As an overall conclusion, the performance of the devices was good and supports their use by farmers for the detection of subclinical ketotic cows in their herds