Friday, April 15, 2016

Whole Blood Gene Expression Profiling in Preclinical and Clinical Cattle Infected with Atypical Bovine Spongiform Encephalopathy

Whole Blood Gene Expression Profiling in Preclinical and Clinical Cattle Infected with Atypical Bovine Spongiform Encephalopathy


Elena Xerxa, Maura Barbisin, Maria Novella Chieppa, Helena Krmac, Elena Vallino Costassa, Paolo Vatta, Marion Simmons, Maria Caramelli, Cristina Casalone, Cristiano Corona, Giuseppe Legname




Published: April 13, 2016 •




Prion diseases, such as bovine spongiform encephalopathies (BSE), are transmissible neurodegenerative disorders affecting humans and a wide variety of mammals. Variant Creutzfeldt-Jakob disease (vCJD), a prion disease in humans, has been linked to exposure to BSE prions. This classical BSE (cBSE) is now rapidly disappearing as a result of appropriate measures to control animal feeding. Besides cBSE, two atypical forms (named H- and L-type BSE) have recently been described in Europe, Japan, and North America. Here we describe the first wide-spectrum microarray analysis in whole blood of atypical BSE-infected cattle. Transcriptome changes in infected animals were analyzed prior to and after the onset of clinical signs. The microarray analysis revealed gene expression changes in blood prior to the appearance of the clinical signs and during the progression of the disease. A set of 32 differentially expressed genes was found to be in common between clinical and preclinical stages and showed a very similar expression pattern in the two phases. A 22-gene signature showed an oscillating pattern of expression, being differentially expressed in the preclinical stage and then going back to control levels in the symptomatic phase. One gene, SEL1L3, was downregulated during the progression of the disease. Most of the studies performed up to date utilized various tissues, which are not suitable for a rapid analysis of infected animals and patients. Our findings suggest the intriguing possibility to take advantage of whole blood RNA transcriptional profiling for the preclinical identification of prion infection. Further, this study highlighted several pathways, such as immune response and metabolism that may play an important role in peripheral prion pathogenesis. Finally, the gene expression changes identified in the present study may be further investigated as a fingerprint for monitoring the progression of disease and for developing targeted therapeutic interventions.




Discussion Whole blood is the most suitable tissue for a prospective rapid diagnostic test since it minimizes sample handling artifacts and reduces sample variability due to fractionation. The present study revealed a substantial gene expression alteration in whole blood from atypical BSE-infected cattle, which could be investigated in future experiments and, if confirmed, could be exploited as a signature for the disease. One of the major caveats in using peripheral blood is that its cellular components may change dramatically during infections or inflammation. The animals used in the present study did not show any apparent side pathology, they were monitored daily by the husbandry staff and their blood was examined for serum aspartate aminotransferase (AST), creatine kinase (CK) and manganese [23]. Nonetheless, possible interference due to hidden pathologies or to inter-individual variations in hematocrit and white blood cell count may affect interpretation of expression data and should be considered as an important variable in future studies. In light of this, the present results should be read as a first exploration of whole blood transcriptomics during a prion infection. To our knowledge, this is the first microarray study of whole blood from BSE-infected cattle. Indeed, in a study published by Panelli et al. [36], fractionated white blood cells were analyzed to detect gene expression changes in L-type infected animals. Very few DEGs are common between the two studies. However, this discrepancy may be explained by different infection methods, microarray platforms, statistical analysis stringency and p value cut-off. Also, the white cells used in the study of Panelli et al. were isolated from 1 year post-infection animals, while in our study we used whole blood from preclinical and clinical infected cattle (around 6 months and 22–26 post-infection respectively).


In the present study, 4 statistical comparisons were performed: infected (preclinical and clinical) versus control (IvsCtrl), preclinical versus control (PvsCtrl), clinical versus control (CvsCtrl) and clinical versus preclinical (CvsP) comparisons. Since our goal was to find a common pattern among all the atypical BSE-infected cattle, we defined them as a single group of infected (H-type and L-type) animals. Indeed, as published by Konold and colleagues [23], these animals shared a very similar phenotype in terms of behavioral and clinical signs. Also, another study by Priemer and colleagues indicated a large similarity at level of PrPSc anatomical distribution for the atypical strains, with only slight differences in the overall intensities between H- and L-type [27]. Nonetheless, even if H- and L-type BSE are reported to share many similarities, they constitute two distinct BSE variants which are characterized by a different electrophoretic mobility of PrPSc unglycosylated moiety after proteinase K (PK) digestion [16, 18]. Statistical comparison between the 4 H-type and the 4 L-type infected animals was carried out in preliminary analyses (see S4 Table), but only a limited number of DEGs was found. Among them, only 15 had a p value lower than 0.01 and only 16 showed a fold change higher than 3, indicating that, at least in terms of number of DEGs, these two groups did not display large differences. For these reasons, we decided to focus our attention on finding a common gene pattern among all the atypical BSE-infected cattle and therefore we pooled the two groups. Due to the high inter-animal variability, which is expected for outbreed animals, further studies in larger animal cohorts would be required to investigate in detail the strain-specific gene expression changes occurring during the progression of the disease. Still, the HvsL analysis can be used as a sort of internal control in this study.


Another aspect to be taken into account when reading the present results is that additional negative control cattle, aged from 12 to 37 months and derived from a different herd compared to the Konold’s study groups [23], were introduced in the analyses. The addition of these controls was useful to balance the samples from infected animals and allowed a preliminary exploration of the differentially expressed transcripts. However, age-related and environmental variability may have affected in some degree the data and need to be considered for their correct interpretation. Despite some limitations, since several statistical analyses were performed (including the CvsP analysis, in which all the animals derived from the same herd) a cross comparison of all them, as we did with the Venn Diagram, may be very useful in order to define a set of genes which could be a good starting point for further validation experiments in the future. In the first statistical analysis we performed (IvsC), we found that among 101 DEGs, 93 had known functions and were involved in several biological processes and molecular pathways, such as autoimmune thyroiditis, chemokine and cytokine activity, regulation of the secretion pathway, the immune system and antigen presentation[52]. Previous studies on CNS tissues from BSE-infected animals also showed the involvement of many of these pathways in prion pathogenesis [4, 35, 52]. This similarity between brain and blood may not be surprising, since it has been shown in the literature that blood transcriptome analyses identify genes that are relevant to the pathological processes occurring in the CNS [53]. Indeed, measuring disease-related gene expression in peripheral blood may be a useful proxy measure for gene expression in the CNS [53, 54].


To characterize the gene expression profile in the preclinical and clinical stages, we performed the PvsCtrl and the CvsCtrl statistical comparisons. We found that 113 probe sets were differentially regulated in the preclinical stage of the disease, while 207 probe sets had an altered expression in the clinical phase. Importantly, the present results indicated that, at least in blood, a consistent gene expression alteration is present from the early stages of the disease. This finding is in agreement with microarray analysis carried out by Tang et al., which revealed the highest degree of differential gene regulation in brains of cBSE-infected cattle at 21 months post infection, which is prior to the detection of infectivity [4]. Also, Tortosa and colleagues found a significant number of DEGs at early stages of the disease in the CNS from cBSE-infected transgenic mice [52].


Venn diagram analysis revealed that 32 DEGs were in common between the clinical and preclinical groups and, remarkably, they had a very similar pattern of expression in both stages of the disease. Since these genes are altered in both phases, it would be very interesting to confirm their differential expression in future experiments with additional negative controls, and eventually in blood from human patients.


Based on GO enrichment analysis, we found that immunity and inflammation processes were strongly involved during the progression of the disease stages. Interestingly, we found that antigen processing and presentation via MHC (major histocompatibility complex) molecules and the autoimmune thyroiditis pathway were significantly altered in atypical BSE-challenged animals. The majority of MHC class I molecule coding-genes were down-regulated in infected cattle (three out of four probes) and, also, MHC class II molecule coding transcripts were found to be down-regulated during the progression of the clinical signs (four out of four probes were down-regulated in the CvsP comparison). The involvement of MHC transcripts in prion pathogenesis is supported by another microarray study published by Khaniya and colleagues in 2009 [55]. In line with the trend found by the microarray analysis, the RT-qPCR validation experiments indicated a downregulation for MHC class I heavy chain (BOLA), even though the results failed to reach the statistical significance (data not shown).


Regarding the autoimmune thyroiditis pathway, it is well known in the literature that Hashimoto’s encephalitis, together with the associated thyroiditis, is a differential diagnosis for CJD, since the two pathologies share a very similar clinical symptomatology [56]. As hypothesized previously by Prusiner and colleagues, the clinical and neuropathological similarities between CJD and Hashimoto’s thyroiditis raise the possibility that protein misprocessing may underlie both neurodegenerative and autoimmune diseases [57].


Finally, a fourth statistical analysis was performed to identify any specific changes between the clinical and the preclinical stages of disease (CvsP). Indeed, we found that the last phases of the disease are accompanied by the overactivation of several genes involved in the immune defense response. In particular, the shift from the preclinical towards the clinical stage was characterized by the upregulation of genes involved in B cell proliferation and the ISG15 (IFN-induced 15-kDa protein) conjugation system. ISG15 is a ubiquitin-like molecule that is tightly regulated by specific innate immunity signaling pathways [58]. Interestingly, it has been shown in the literature that this protein is over-activated in the spinal cord of amyotrophic lateral sclerosis mice models [59] and it has been indicated as a general marker for both acute and chronic neuronal injuries [60].


To further analyze the data, we compared the list of DEGs found in PvsCtrl, CvsP and CvsCtrl and found 22 genes with an oscillatory pattern of expression, being differentially expressed in the preclinical stage and then going back roughly to the control level in the clinical stage. Interestingly, some of the oscillatory DEGs are involved in regulation of transcription, thus suggesting that the gene expression during atypical BSE infection is tightly regulated. Venn diagram analysis revealed that one gene, SEL1L3, was down-regulated in all the comparisons (PvsCtrl, CvsCtrl, CvsP). SEL1L3 codes for a transmembrane protein whose function is unknown. Interestingly, an important paralog of SEL1L3, SEL1L, is involved in the retrotranslocation of misfolded proteins from the lumen of the endoplasmic reticulum to the cytosol, where they are degraded by the proteasome in an ubiquitin-dependent manner [61]. Therefore, we could hypothesize that its down-regulation in prion infected animals would lead to a reduced degradation of PrPSc, thus supporting the progression of the disease. We validated this gene by RT-qPCR, confirming its downregulation in both the preclinical and clinical stages of the disease. Further investigation on the function of SEL1L3L would be of great interest since this gene may play an important role in prion disease and maybe other neurodegenerative illnesses.


Besides SEL1L3, five other genes were validated by RT-qPCR; here we will briefly discuss how these genes may be involved in prion pathogenesis and in host response to prion infection.


GNLY and CD40L were found to be down-regulated in both preclinical and clinical stages. GNLY is a powerful antimicrobial protein contained within the granules of cytotoxic T lymphocyte and natural killer cells. This gene was found to be downregulated also in a microarray study performed on the medulla oblongata from sheep with preclinical natural scrapie [62]. Thus, it may be a good candidate as an early biomarker for atypical BSE but also for other prion diseases.


CD40–CD40L interactions mediate a broad variety of immune and inflammatory responses and have been implicated in the pathogenesis of Alzheimer’s disease (AD) [63, 64]. Although the importance of CD40L in prion disease progression has not yet been clarified (66–68), its downregulation in blood during both preclinical and clinical stages of atypical BSE-infection suggests that prion infection has an impact on the host immune system response and that immune tolerance may be an active process induced by prions.


Two other downregulated genes were validated by RT-qPCR, namely HBA2 and XIST. Concerning HBA2, we found a downregulation in preclinical atypical BSE-infected cattle. Haemoglobins are iron-containing proteins that transport oxygen in the blood of most vertebrates. Beside blood, HBA and HBB are also expressed in mesencephalic dopaminergic neurons and glial cells [65] and are down regulated in AD, PD and other neurodegenerative diseases [66]. Haemoglobin genes expression alteration during preclinical scrapie was also found in the spleen and CNS of infected animals [67, 68], as well as in the brains of nonhuman primates infected with BSE [51]. These findings suggest an involvement of these genes in the host response to general neurodegenerative processes. Besides changes in transcript levels, it has been found that both HbA and HbB protein distribution is altered in mitochondrial fractions from PD degenerating brain [69]. Moreover, HbA is also expressed in endothelial cells, where it regulates the nitric oxide signaling [70]. Even though a clear mechanism linking these molecules to neurodegeneration has not yet been described, taken together these findings strongly suggest a central role for haemoglobin in neurodegenerative processes.


A marked downregulation in XIST expression was found in our infected animals. XIST is a gene located on X chromosomes which codes for a long non-coding RNA (LncRNA) involved in X-chromosome dosage control [71, 72]. LncRNAs are emerging as useful biomarkers for neurodegenerative diseases such as AD [73] and other disease processes [74], and they can be easily detected in blood and urine from patients. In addition, we cannot exclude the possibility that the alteration in XIST expression may have some role in gender-dependent response to prion infection [75].


RT-qPCR experiments confirmed the upregulation of PDK4 in clinically affected animals. PDK4 encodes for a mitochondrial protein involved in glucose metabolism through the inhibition of pyruvate dehydrogenase complex, which leads to a reduction in pyruvate conversion to acetyl-CoA [76]. In the literature, a key role has been suggested for acetyl-CoA fueling for the survival of cholinergic neurons in the course of neurodegenerative diseases [77]. PDK4 overactivation can lead to a switch from glucose catabolism to fatty acid utilization [78], thus increasing the production of ketone bodies. Notably, it has been shown in the literature that these molecules are able to cross the blood brain barrier. We could speculate that in prion infection (or at least in atypical BSE infection) the concentration of ketone bodies would rise in blood, as a consequence of PDK4 upregulation, and act in the brain as neuroprotective molecules [79, 80]. This would be an attempt by the organism to prevent the neurodegeneration induced by prions.




 Conclusions In conclusion, the present study has led to the identification of several gene expression changes in whole blood from clinical and preclinical atypical BSE cattle, which upon further investigation and validation in blood from human patients, might represent a molecular fingerprint to characterize this disease. By comparing our results with other studies on various animal prion diseases, we observed that some of the most significantly altered DEGs we found in blood were found differentially expressed also in brain tissue from BSE-infected cattle; this observation indicates that whole blood transcriptome analyses may serve as a proxy measure for the changes occurring in the CNS of infected animals. Furthermore, our study underlines the importance of utilizing whole blood, without any additional manipulation, as a source tissue as it is an easily accessible body fluid. In addition, the transcription regulation activated in atypical BSE infections is similar to some extent to the one observed in the literature for cBSE, even though the clinical characteristics and biochemical properties are very different. Thus, this gene expression profile may be investigated in other BSE infections to identify a common molecular fingerprint.


Overall, our study confirmed the differential expression of 6 genes (XIST, CD40L, GNLY, PDK4, HBA2 and SEL1L3), which may play several roles in atypical BSE pathogenesis and, possibly, in other prion infections. Indeed, they are involved in multiple pathways such as immune response, inflammation, and glucose catabolism. Even though further studies are required to investigate the specific involvement of all the identified genes in prion diseases, our data indicate an important role for immune system regulation in the prion pathogenesis of atypical BSE and maybe in BSE as well as in other prion diseases in general.



Wednesday, September 23, 2015


NIH Availability for Licensing AGENCY: [FR Doc. 2015–24117 Filed 9–22–15; 8:45 am] Detection and Discrimination of Classical and Atypical L-Type BSE Strains by RT-QuIC



Monday, March 19, 2012


Infectivity in Skeletal Muscle of Cattle with Atypical Bovine Spongiform Encephalopathy PLoS One. 2012; 7(2): e31449.





***********OCTOBER 2015*************




THANK YOU PRION 2015 TAYLOR & FRANCIS, Professor Chernoff, and Professor Aguzzi et al, for making these PRION 2015 Congressional Poster and Oral Abstracts available freely to the public. ...Terry S. Singeltary Sr.


O.05: Transmission of prions to primates after extended silent incubation periods: Implications for BSE and scrapie risk assessment in human populations


Emmanuel Comoy, Jacqueline Mikol, Val erie Durand, Sophie Luccantoni, Evelyne Correia, Nathalie Lescoutra, Capucine Dehen, and Jean-Philippe Deslys Atomic Energy Commission; Fontenay-aux-Roses, France


Prion diseases (PD) are the unique neurodegenerative proteinopathies reputed to be transmissible under field conditions since decades. The transmission of Bovine Spongiform Encephalopathy (BSE) to humans evidenced that an animal PD might be zoonotic under appropriate conditions. Contrarily, in the absence of obvious (epidemiological or experimental) elements supporting a transmission or genetic predispositions, PD, like the other proteinopathies, are reputed to occur spontaneously (atpical animal prion strains, sporadic CJD summing 80% of human prion cases). Non-human primate models provided the first evidences supporting the transmissibiity of human prion strains and the zoonotic potential of BSE. Among them, cynomolgus macaques brought major information for BSE risk assessment for human health (Chen, 2014), according to their phylogenetic proximity to humans and extended lifetime. We used this model to assess the zoonotic potential of other animal PD from bovine, ovine and cervid origins even after very long silent incubation periods. ***We recently observed the direct transmission of a natural classical scrapie isolate to macaque after a 10-year silent incubation period, with features similar to some reported for human cases of sporadic CJD, albeit requiring fourfold longe incubation than BSE. ***Scrapie, as recently evoked in humanized mice (Cassard, 2014), is the third potentially zoonotic PD (with BSE and L-type BSE), ***thus questioning the origin of human sporadic cases. We will present an updated panorama of our different transmission studies and discuss the implications of such extended incubation periods on risk assessment of animal PD for human health.




***thus questioning the origin of human sporadic cases...





***Our study demonstrates susceptibility of adult cattle to oral transmission of classical BSE. ***


***our findings suggest that possible transmission risk of H-type BSE to sheep and human. ***


P.86: Estimating the risk of transmission of BSE and scrapie to ruminants and humans by protein misfolding cyclic amplification


Morikazu Imamura, Naoko Tabeta, Yoshifumi Iwamaru, and Yuichi Murayama National Institute of Animal Health; Tsukuba, Japan


To assess the risk of the transmission of ruminant prions to ruminants and humans at the molecular level, we investigated the ability of abnormal prion protein (PrPSc) of typical and atypical BSEs (L-type and H-type) and typical scrapie to convert normal prion protein (PrPC) from bovine, ovine, and human to proteinase K-resistant PrPSc-like form (PrPres) using serial protein misfolding cyclic amplification (PMCA).


Six rounds of serial PMCA was performed using 10% brain homogenates from transgenic mice expressing bovine, ovine or human PrPC in combination with PrPSc seed from typical and atypical BSE- or typical scrapie-infected brain homogenates from native host species. In the conventional PMCA, the conversion of PrPC to PrPres was observed only when the species of PrPC source and PrPSc seed matched. However, in the PMCA with supplements (digitonin, synthetic polyA and heparin), both bovine and ovine PrPC were converted by PrPSc from all tested prion strains. On the other hand, human PrPC was converted by PrPSc from typical and H-type BSE in this PMCA condition.


Although these results were not compatible with the previous reports describing the lack of transmissibility of H-type BSE to ovine and human transgenic mice, ***our findings suggest that possible transmission risk of H-type BSE to sheep and human. Bioassay will be required to determine whether the PMCA products are infectious to these animals.








***our findings suggest that possible transmission risk of H-type BSE to sheep and human. Bioassay will be required to determine whether the PMCA products are infectious to these animals.






Title: Transmission of scrapie prions to primate after an extended silent incubation period




item Comoy, Emmanuel - item Mikol, Jacqueline - item Luccantoni-Freire, Sophie - item Correia, Evelyne - item Lescoutra-Etchegaray, Nathalie - item Durand, Valérie - item Dehen, Capucine - item Andreoletti, Olivier - item Casalone, Cristina - item Richt, Juergen item Greenlee, Justin item Baron, Thierry - item Benestad, Sylvie - item Hills, Bob - item Brown, Paul - item Deslys, Jean-Philippe -


Submitted to: Scientific Reports Publication Type: Peer Reviewed Journal Publication Acceptance Date: May 28, 2015 Publication Date: June 30, 2015 Citation: Comoy, E.E., Mikol, J., Luccantoni-Freire, S., Correia, E., Lescoutra-Etchegaray, N., Durand, V., Dehen, C., Andreoletti, O., Casalone, C., Richt, J.A., Greenlee, J.J., Baron, T., Benestad, S., Brown, P., Deslys, J. 2015. Transmission of scrapie prions to primate after an extended silent incubation period. Scientific Reports. 5:11573.


Interpretive Summary:


The transmissible spongiform encephalopathies (also called prion diseases) are fatal neurodegenerative diseases that affect animals and humans. The agent of prion diseases is a misfolded form of the prion protein that is resistant to breakdown by the host cells. Since all mammals express prion protein on the surface of various cells such as neurons, all mammals are, in theory, capable of replicating prion diseases. One example of a prion disease, bovine spongiform encephalopathy (BSE; also called mad cow disease), has been shown to infect cattle, sheep, exotic undulates, cats, non-human primates, and humans when the new host is exposed to feeds or foods contaminated with the disease agent. The purpose of this study was to test whether non-human primates (cynomologous macaque) are susceptible to the agent of sheep scrapie. After an incubation period of approximately 10 years a macaque developed progressive clinical signs suggestive of neurologic disease. Upon postmortem examination and microscopic examination of tissues, there was a widespread distribution of lesions consistent with a transmissible spongiform encephalopathy. This information will have a scientific impact since it is the first study that demonstrates the transmission of scrapie to a non-human primate with a close genetic relationship to humans. This information is especially useful to regulatory officials and those involved with risk assessment of the potential transmission of animal prion diseases to humans.


Technical Abstract:


Classical bovine spongiform encephalopathy (c-BSE) is an animal prion disease that also causes variant Creutzfeldt-Jakob disease in humans. Over the past decades, c-BSE's zoonotic potential has been the driving force in establishing extensive protective measures for animal and human health. In complement to the recent demonstration that humanized mice are susceptible to scrapie, we report here the first observation of direct transmission of a natural classical scrapie isolate to a macaque after a 10-year incubation period. Neuropathologic examination revealed all of the features of a prion disease: spongiform change, neuronal loss, and accumulation of PrPres throughout the CNS.


***This observation strengthens the questioning of the harmlessness of scrapie to humans, at a time when protective measures for human and animal health are being dismantled and reduced as c-BSE is considered controlled and being eradicated. Our results underscore the importance of precautionary and protective measures and the necessity for long-term experimental transmission studies to assess the zoonotic potential of other animal prion strains.



***This information will have a scientific impact since it is the first study that demonstrates the transmission of scrapie to a non-human primate with a close genetic relationship to humans. This information is especially useful to regulatory officials and those involved with risk assessment of the potential transmission of animal prion diseases to humans.


***This observation strengthens the questioning of the harmlessness of scrapie to humans, at a time when protective measures for human and animal health are being dismantled and reduced as c-BSE is considered controlled and being eradicated. Our results underscore the importance of precautionary and protective measures and the necessity for long-term experimental transmission studies to assess the zoonotic potential of other animal prion strains.









Zoonotic Potential of CWD Prions


Liuting Qing1, Ignazio Cali1,2, Jue Yuan1, Shenghai Huang3, Diane Kofskey1, Pierluigi Gambetti1, Wenquan Zou1, Qingzhong Kong1 1Case Western Reserve University, Cleveland, Ohio, USA, 2Second University of Naples, Naples, Italy, 3Encore Health Resources, Houston, Texas, USA


*** These results indicate that the CWD prion has the potential to infect human CNS and peripheral lymphoid tissues and that there might be asymptomatic human carriers of CWD infection.




***These results indicate that the CWD prion has the potential to infect human CNS and peripheral lymphoid tissues and that there might be asymptomatic human carriers of CWD infection.***




P.105: RT-QuIC models trans-species prion transmission


Kristen Davenport, Davin Henderson, Candace Mathiason, and Edward Hoover Prion Research Center; Colorado State University; Fort Collins, CO USA


Conversely, FSE maintained sufficient BSE characteristics to more efficiently convert bovine rPrP than feline rPrP. Additionally, human rPrP was competent for conversion by CWD and fCWD.


***This insinuates that, at the level of protein:protein interactions, the barrier preventing transmission of CWD to humans is less robust than previously estimated.




***This insinuates that, at the level of protein:protein interactions, the barrier preventing transmission of CWD to humans is less robust than previously estimated.***







Transmissible Spongiform Encephalopathy TSE PRION update January 2, 2014


*** chronic wasting disease, there was no absolute barrier to conversion of the human prion protein.


*** Furthermore, the form of human PrPres produced in this in vitro assay when seeded with CWD, resembles that found in the most common human prion disease, namely sCJD of the MM1 subtype.




*** These results would seem to suggest that CWD does indeed have zoonotic potential, at least as judged by the compatibility of CWD prions and their human PrPC target. Furthermore, extrapolation from this simple in vitro assay suggests that if zoonotic CWD occurred, it would most likely effect those of the PRNP codon 129-MM genotype and that the PrPres type would be similar to that found in the most common subtype of sCJD (MM1).***



*** The potential impact of prion diseases on human health was greatly magnified by the recognition that interspecies transfer of BSE to humans by beef ingestion resulted in vCJD. While changes in animal feed constituents and slaughter practices appear to have curtailed vCJD, there is concern that CWD of free-ranging deer and elk in the U.S. might also cross the species barrier. Thus, consuming venison could be a source of human prion disease. Whether BSE and CWD represent interspecies scrapie transfer or are newly arisen prion diseases is unknown. Therefore, the possibility of transmission of prion disease through other food animals cannot be ruled out. There is evidence that vCJD can be transmitted through blood transfusion. There is likely a pool of unknown size of asymptomatic individuals infected with vCJD, and there may be asymptomatic individuals infected with the CWD equivalent. These circumstances represent a potential threat to blood, blood products, and plasma supplies.



Tuesday, April 12, 2016


The first detection of Chronic Wasting Disease (CWD) in Europe Norway



Thursday, January 29, 2015


Atypical H-TYPE BSE Case Confirmed in Norway



Monday, April 11, 2016







Friday, February 20, 2015


***APHIS Freedom of Information Act (FOIA) Appeal Mouse Bio-Assays 2007-00030-A Sheep Imported From Belgium and the Presence of TSE Prion Disease Kevin Shea to Singeltary 2015



Friday, March 18, 2016 CFSAN


Constituent Update: FDA Announces Final Rule on Bovine Spongiform Encephalopathy BSE MAD COW TSE PRION Center for Food Safety and Applied Nutrition - Constituent Update



Tuesday, March 15, 2016


*** Docket No. FDA-2016-N-0321 Risk Assessment of Foodborne Illness Associated with Pathogens from Produce Grown in Fields Amended with Untreated Biological Soil Amendments of Animal Origin; Request for Comments, Scientific Data, and Information Singeltary Submission ***



Monday, March 28, 2016


National Scrapie Eradication Program February 2016 Monthly Report



*** Docket No. APHIS-2007-0127 Scrapie in Sheep and Goats Terry Singeltary Sr. Submission ***


Monday, November 16, 2015


*** Docket No. APHIS-2007-0127 Scrapie in Sheep and Goats Terry Singeltary Sr. Submission ***




Saturday, July 18, 2015






*** We describe the transmission of spongiform encephalopathy in a non-human primate inoculated 10 years earlier with a strain of sheep c-scrapie. Because of this extended incubation period in a facility in which other prion diseases are under study, we are obliged to consider two alternative possibilities that might explain its occurrence.


We first considered the possibility of a sporadic origin (like CJD in humans). Such an event is extremely improbable because the inoculated animal was 14 years old when the clinical signs appeared, i.e. about 40% through the expected natural lifetime of this species, compared to a peak age incidence of 60–65 years in human sporadic CJD, or about 80% through their expected lifetimes.


***Moreover, sporadic disease has never been observed in breeding colonies or primate research laboratories, most notably among hundreds of animals over several decades of study at the National Institutes of Health25, and in nearly twenty older animals continuously housed in our own facility.***


Epidemiology | Neurological manifestations | Prion diseases Received: 16 February 2015 Accepted: 28 May 2015 Published online: 30 June 2015


Transmission of scrapie prions to primate after an extended silent incubation period





 Thursday, March 24, 2016





Sunday, October 5, 2014


France stops BSE testing for Mad Cow Disease



***atypical spontaneous BSE in France LOL***


FRANCE STOPS TESTING FOR MAD COW DISEASE BSE, and here’s why, to many spontaneous events of mad cow disease $$$


If you Compare France to other Countries with atypical BSE, in my opinion, you cannot explain this with ‘spontaneous’.


Table 1: Number of Atypical BSE cases reported by EU Member States in the period 2001–2014 by country and by type (L- and H-BSE) (extracted from EU BSE databases on 1 July 2014). By 2015, these data might be more comprehensive following a request from the European Commission to Member States for re-testing and retrospective classification of all positive bovine isolates in the EU in the years 2003–2009


BSE type


Country 2001 2002 2003 2004 2005 2006 2007 2008 2009 2010 2011 2012 2013(a) 2014(a) Total


H-BSE Austria 1 1


France(b) 1 2 3 1 2 2 2 2 15


Germany 1 1 2


Ireland 1 1 2 1 5


The Netherlands 1 1


Poland 1 1 2


Portugal 1 1


Spain 1 1 2


Sweden 1 1


United Kingdom 1 1 1 1 1 5


Total 2 3 3 1 1 2 2 2 4 4 5 1 4 1 35


L-BSE Austria 1 1 2


Denmark 1 1


France(b) 1 1 1 1 2 1 3 2 1 1 14


Germany 1 1 2


Italy 1 1 1 1 1 5


The Netherlands 1 1 1 3


Poland 1 2 2 1 2 1 2 1 12


Spain 2 2


United Kingdom 1 1 1 1 4


Total 0 5 3 4 3 3 6 3 3 4 3 6 1 1 45


Total Atypical cases (H + L)


2 8 6 5 4 5 8 5 7 8 8 7 5 2 80


(a): Data for 2013-2014 are incomplete and may not include all cases/countries reported.


(b): France has performed extensive retrospective testing to classify BSE cases, which is probably the explanation for the higher number of Atypical BSE cases reported in this country.


The number of Atypical BSE cases detected in countries that have already identified them seems to be similar from year to year. In France, a retrospective study of all TSE-positive cattle identified through the compulsory EU surveillance between 2001 and 2007 indicated that the prevalence of H-BSE and L-BSE was 0.35 and 0.41 cases per million adult cattle tested, respectively, which increased to 1.9 and 1.7 cases per million, respectively, in tested animals over eight years old (Biacabe et al., 2008). No comprehensive study on the prevalence of Atypical BSE cases has yet been carried out in other EU Member States. All cases of Atypical BSE reported in the EU BSE databases have been identified by active surveillance testing (59 % in fallen stock, 38 % in healthy slaughtered cattle and 4 % in emergency slaughtered cattle). Cases were reported in animals over eight years of age, with the exception of two cases (one H-BSE and one L-BSE) detected in Spain in 2011/2012. One additional case of H-BSE was detected in Switzerland in 2012 in a cow born in Germany in 2005 (Guldimann et al., 2012).




Wednesday, July 15, 2015


Additional BSE TSE prion testing detects pathologic lesion in unusual brain location and PrPsc by PMCA only, how many cases have we missed?





Sunday, March 20, 2016


Docket No. FDA-2003-D-0432 (formerly 03D-0186) Use of Material from Deer and Elk in Animal Feed ***UPDATED MARCH 2016*** Singeltary Submission



Friday, February 05, 2016


*** Report of the Committee on Wildlife Diseases FY2015 CWD TSE PRION Detections in Farmed Cervids and Wild



Saturday, February 6, 2016


*** Secretary's Advisory Committee on Animal Health; Meeting [Docket No. APHIS-2016-0007] Singeltary Submission



Thursday, January 14, 2016


EMERGING ANIMAL DISEASES Actions Needed to Better Position USDA to Address Future Risks Report to the Chairman, Committee on Energy and Commerce, House of Representatives December 2015 GAO-16-132





Saturday, January 9, 2016


Transmission of sheep-bovine spongiform encephalopathy to pigs


Research article



Long live the OIE, or time to close the doors on a failed entity?


Long live the OIE


By Dr. Bernard Vallat, OIE Director General December 29, 2015 | 10:08 am EST



Wednesday, April 25, 2012





Saturday, January 29, 2011


Atypical L-Type Bovine Spongiform Encephalopathy (L-BSE) Transmission to Cynomolgus Macaques, a Non-Human Primate




Monday, September 13, 2010


atypical BSE strains and sporadic CJD strains, is there a connection and why shouldn't there be $



Sunday, April 12, 2009





Wednesday, March 26, 2008


MAD COW DISEASE terminology UK c-BSE (typical), atypical BSE H or L, and or Italian L-BASE



Thursday, January 3, 2008





Thursday, April 14, 2016


Arizona 22 year old diagnosed with Creutzfeldt Jakob Disease CJD



Saturday, January 16, 2016


Revised Preventive Measures to Reduce the Possible Risk of Transmission of Creutzfeldt-Jakob Disease and Variant Creutzfeldt-Jakob Disease by Blood and Blood Products Guidance for Industry



Thursday, September 10, 2015


25th Meeting of the Transmissible Spongiform Encephalopathies Advisory Committee Food and Drug Administration Silver Spring, Maryland June 1, 2015




Terry S. Singeltary Sr.

Saturday, May 09, 2015

Expression of genes involved in the T cell signalling pathway in circulating immune cells of cattle 24 months following oral challenge with Bovine Amyloidotic Spongiform Encephalopathy (BASE)

Research article

Expression of genes involved in the T cell signalling pathway in circulating immune cells of cattle 24 months following oral challenge with Bovine Amyloidotic Spongiform Encephalopathy (BASE)

Andrea Trovato1, Simona Panelli2, Francesco Strozzi1, Caterina Cambulli2, Ilaria Barbieri3, Nicola Martinelli3, Guerino Lombardi3, Rossana Capoferri2 and John L Williams14*

* Corresponding author: John L Williams

Author Affiliations

1 Parco Tecnologico Padano, via Einstein, Lodi 26900, Italy

2 Istituto Sperimentale Italiano Lazzaro Spallanzani, Loc. La Quercia, Rivolta d’Adda, 26027, Italy

3 Istituto Zooprofilattico Sperimentale della Lombardia e dell’Emilia Romagna, via Bianchi 9, Brescia, 25124, Italy

4 Present address: School of Animal and Veterinary Sciences, University of Adelaide, Roseworthy 5371, SA, Australia

For all author emails, please log on.

BMC Veterinary Research 2015, 11:105 doi:10.1186/s12917-015-0412-y

Published: 9 May 2015


Background Bovine Amyloidotic Spongiform Encephalopathy (BASE) is a variant of classical BSE that affects cows and can be transmitted to primates and mice. BASE is biochemically different from BSE and shares some molecular and histo-pathological features with the MV2 sub-type of human sporadic Creutzfeld Jakob Disease (sCJD).

Results The present work examined the effects of BASE on gene expression in circulating immune cells. Ontology analysis of genes differentially expressed between cattle orally challenged with brain homogenate from cattle following intracranial inoculation with BASE and control cattle identified three main pathways which were affected. Within the immune function pathway, the most affected genes were related to the T cell receptor-mediated T cell activation pathways. The differential expression of these genes in BASE challenged animals at 10,12 and 24 months following challenge, vs unchallenged controls, was investigated by real time PCR.

Conclusions The results of this study show that the effects of prion diseases are not limited to the CNS, but involve the immune system and particularly T cell signalling during the early stage following challenge, before the appearance of clinical signs.

Keywords: Bovine Amyloidotic Spongiform Encephalopathy (BASE); Transmissible Spongiform Encephalopathies (TSEs); Immune function; Cattle


All the cases of BSE identified during the major outbreak in the UK were of the same strain type [19]. However, an atypical form of BSE, Bovine Amyloidotic Spongiform Encephalopathy (BASE), was discovered in Italy in 2004 in two old (11 and 15 year old) asymptomatic cows post mortem [19]. Other atypical forms of BSE were subsequently reported in France, Germany and Japan [19-22]. The frequency of atypical BSE may be similar to the occurrence of sporadic CJD, which is about 1 per million individuals [23]. BASE can be biochemically differentiated from BSE by the different mobility of PrP fragments on gel electrophoresis. BASE can also be distinguished from BSE histo-pathologically based on differences in the distribution of vacuoles in the brain. ***BASE shares molecular and histopathological features with the MV2 sub-type of human sporadic Creutzfeldt-Jakob Disease (sCJD).



The data presented here on gene expression in circulating immune cells following BASE challenge show that response to BASE has similarities with other prion diseases. PrPc is known to have a role in the immune system, indeed it is expressed on DC and is important for inducing the T cell proliferative response [30]. Moreover, PrPc accumulates in the contact point between T cells and DC, and it may have a role in the assembly of the TCR complex [45]. The disease form of this protein (PrPres) has been shown to affect the immune system, e.g. eliciting qualitative differences in the responses of T cells [46]. Moreover, macrophages accumulate PrPres, and may be involved in the transfer of the disease to the CNS [9,47]. The data presented here are consistent with the hypothesis that the effects of TSE diseases are not limited to CNS, but involve the immune system, especially during the early stages following challenge, before the appearance of clinical signs. Our data suggest that BASE challenge affects the TCR signalling pathway, which has also been shown in mouse knock-out experiments [17]. BASE therefore, in common with other prion diseases, seems to be associated with general cellular stress and impaired immune function. These data, from experimentally challenged cattle, suggest that orally administered BASE affects gene expression in circulating immune cells even in the absence of overt disease.

spontaneous atypical BSE ???

if that's the case, then France is having one hell of an epidemic of atypical BSE, probably why they stopped testing for BSE, problem solved $$$

As of December 2011, around 60 atypical BSE cases have currently been reported in 13 countries, *** with over one third in France.

FRANCE STOPS TESTING FOR MAD COW DISEASE BSE, and here’s why, to many spontaneous events of mad cow disease $$$

so 20 cases of atypical BSE in France, compared to the remaining 40 cases in the remaining 12 Countries, divided by the remaining 12 Countries, about 3+ cases per country, besides Frances 20 cases. you cannot explain this away with any spontaneous BSe. ...TSS

Sunday, October 5, 2014

France stops BSE testing for Mad Cow Disease

Identification of a second bovine amyloidotic spongiform encephalopathy: Molecular similarities with sporadic Creutzfeldt–Jakob disease

Cristina Casalone*†, Gianluigi Zanusso†‡, Pierluigi Acutis*, Sergio Ferrari‡, Lorenzo Capucci§, Fabrizio Tagliavini¶, Salvatore Monaco‡ , and Maria Caramelli* *Centro di Referenza Nazionale per le Encefalopatie Animali, Istituto Zooprofilattico Sperimentale del Piemonte, Liguria e Valle d’Aosta, Via Bologna, 148, 10195 Turin, Italy; ‡Department of Neurological and Visual Science, Section of Clinical Neurology, Policlinico G.B. Rossi, Piazzale L.A. Scuro, 10, 37134 Verona, Italy; §Istituto Zooprofilattico Sperimentale della Lombardia ed Emilia Romagna, Via Bianchi, 9, 25124 Brescia, Italy; and ¶Istituto Nazionale Neurologico ‘‘Carlo Besta,’’ Via Celoria 11, 20133 Milan, Italy Edited by Stanley B. Prusiner, University of California, San Francisco, CA, and approved December 23, 2003 (received for review September 9, 2003)

Transmissible spongiform encephalopathies (TSEs), or prion diseases, are mammalian neurodegenerative disorders characterized by a posttranslational conversion and brain accumulation of an insoluble, protease-resistant isoform (PrPSc) of the host-encoded cellular prion protein (PrPC). Human and animal TSE agents exist as different phenotypes that can be biochemically differentiated on the basis of the molecular mass of the protease-resistant PrPSc fragments and the degree of glycosylation. Epidemiological, molecular, and transmission studies strongly suggest that the single strain of agent responsible for bovine spongiform encephalopathy (BSE) has infected humans, causing variant Creutzfeldt–Jakob disease. The unprecedented biological properties of the BSE agent, which circumvents the so-called ‘‘species barrier’’ between cattle and humans and adapts to different mammalian species, has raised considerable concern for human health. To date, it is unknown whether more than one strain might be responsible for cattle TSE or whether the BSE agent undergoes phenotypic variation after natural transmission. Here we provide evidence of a second cattle TSE. The disorder was pathologically characterized by the presence of PrP-immunopositive amyloid plaques, as opposed to the lack of amyloid deposition in typical BSE cases, and by a different pattern of regional distribution and topology of brain PrPSc accumulation. In addition, Western blot analysis showed a PrPSc type with predominance of the low molecular mass glycoform and a protease- resistant fragment of lower molecular mass than BSE-PrPSc. Strikingly, the molecular signature of this previously undescribed bovine PrPSc was similar to that encountered in a distinct subtype of sporadic Creutzfeldt–Jakob disease.



In natural and experimental TSEs, PrPSc deposition represents an early event that occurs weeks to months before the development of spongiform changes (20, 21). As a consequence, the detection of PrPSc by Western immunoblot provides a unique opportunity in the diagnosis of BSE early in the incubation period and, therefore, in presymptomatic animals. The identification of the present cattle by postmortem biochemical tests, in the absence of clear neurological involvement, suggests that the disorder was detected at early stages, and this may also explain the lack of widespread vacuolar changes.

Previous pathological studies in clinically suspect cases of BSE in Great Britain have provided evidence for a uniform pattern in the severity and distribution of vacuolar lesions in affected animals, with medulla oblongata nuclei being the most involved (22). While confirming that the BSE epidemic has been sustained by a single agent, these studies have assessed the validity of statutory criteria for the diagnosis of BSE, which is currently based on both histopathological and immunobiochemical exam- ination of the medulla. However, the prevailing involvement of cortical regions in the cattle with amyloid deposition suggests that postmortem brain sampling should not be limited to the obex. In addition, a careful analysis of PrPSc glycoform profiles at the confirmatory Western immunoblot may provide a molecular means of identifying atypical cases of bovine TSE.

Bovine Amyloidotic Spongiform Encephalopathy (BASE): A Second Bovine TSE. The present findings show that a previously undescribed pathological and immunohistochemical phenotype, associated with cattle TSE, is related to the presence of a PrPSc type with biochemical properties, including the gel mobility of the protease-resistant fragment and glycoform ratios, different from those encountered in cattle BSE. Brain deposition of this pathological isoform of cattle PrP correlates with the formation of PrP-amyloid plaques, as opposed to typical BSE cases. Although in several natural and experimental recipients of the BSE agent, including humans (13), neuropathological changes are characterized by the presence of PrP-positive amyloid deposits with surrounding vacuolation, cattle BSE is not associated with PrP-amyloid plaque formation. On the basis of the above features, we propose to name the disease described here BASE. Although observed in only two cattle, the BASE phenotype could be more common than expected. In previous studies, amyloid congophilic plaques were found in 1 of 20 BSE cases examined systematically for amyloid (23), and it was reported that focal cerebral amyloidosis is present in a small proportion of BSE cases (24). Although no biochemical analysis of PrPSc glycotype is available for these animals with ‘‘atypical BSE phenotype,’’ our present results underscore the importance of performing a strain-typing in bovine TSE with amyloid deposition.

In sCJD, the neuropathological phenotype largely correlates with the molecular type of PrPSc and distinct polymorphic sites of PRNP (9, 19). This is in contrast with the situation in cattle, where different genotypes have been reported based on the variable numbers of octapeptide repeats in each allele, but no evidence for single-codon polymorphisms in the PrP gene has been established (25, 26). Because the present animals shared a similar genetic background and breed, differences in disease phenotypes between cattle with BSE and BASE can be tentatively related only to distinct PrPSc types or alternative routes of infection and spread of prion pathology. Accordingly, the lack of involvement of the motor dorsal nucleus of the vagus and the slight involvement of the brainstem in BASE, suggests a route for spreading of the agent other than the alimentary tract. Therefore, unless the BASE agent propagates throughout the olfactory pathway or other peripheral routes, it is possible that this disorder represents a sporadic form of cattle TSE, which would also explain the difference in ages between the two groups of affected animals.

Phenotypic Similarities Between BASE and sCJD. The transmissibility of CJD brains was initially demonstrated in primates (27), and classification of atypical cases as CJD was based on this property (28). To date, no systematic studies of strain typing in sCJD have been provided, and classification of different subtypes is based on clinical, neuropathological, and molecular features (the polymorphic PRNP codon 129 and the PrPSc glycotype) (8, 9, 15, 19).

The importance of molecular PrPSc characterization in assessing the identity of TSE strains is underscored by several studies, showing that the stability of given disease-specific PrPSc types is maintained upon experimental propagation of sCJD, familial CJD, and vCJD isolates in transgenic PrP-humanized mice (8, 29). Similarly, biochemical properties of BSE- and vCJDassociated PrPSc molecules remain stable after passage to mice expressing bovine PrP (30). Recently, however, it has been reported that PrP-humanized mice inoculated with BSE tissues may also propagate a distinctive PrPSc type, with a ‘‘monoglycosylated- dominant’’ pattern and electrophoretic mobility of the unglycosylated fragment slower than that of vCJD and BSE (31). Strikingly, this PrPSc type shares its molecular properties with the a PrPSc molecule found in classical sCJD. This observation is at variance with the PrPSc type found in M V2 sCJD cases and in cattle BASE, showing a monoglycosylated-dominant pattern but faster electrophoretic mobility of the protease-resistant fragment as compared with BSE. In addition to molecular properties of PrPSc, BASE and M V2 sCJD share a distinctive pattern of intracerebral PrP deposition, which occurs as plaque-like and amyloid-kuru plaques. Differences were, however, observed in the regional distribution of PrPSc. While inM V2 sCJD cases the largest amounts of PrPSc were detected in the cerebellum, brainstem, and striatum, in cattle BASE these areas were less involved and the highest levels of PrPSc were recovered from the thalamus and olfactory regions.

In conclusion, decoding the biochemical PrPSc signature of individual human and animal TSE strains may allow the identification of potential risk factors for human disorders with unknown etiology, such as sCJD. However, although BASE and sCJD share several characteristics, caution is dictated in assessing a link between conditions affecting two different mammalian species, based on convergent biochemical properties of diseaseassociated PrPSc types. Strains of TSE agents may be better characterized upon passage to transgenic mice. In the interim until this is accomplished, our present findings suggest a strict epidemiological surveillance of cattle TSE and sCJD based on molecular criteria.



Summary Report BSE 2012

Executive Summary

Saturday, August 4, 2012

 *** Final Feed Investigation Summary - California BSE Case - July 2012 ***

Saturday, August 4, 2012

 Update from APHIS Regarding Release of the Final Report on the BSE Epidemiological Investigation

Atypical BSE (BASE) Transmitted from Asymptomatic Aging Cattle to a Primate

Conclusion/Significance Our results point to a possibly higher degree of pathogenicity of BASE than classical BSE in primates and also raise a question about a possible link to one uncommon subset of cases of apparently sporadic CJD. Thus, despite the waning epidemic of classical BSE, the occurrence of atypical strains should temper the urge to relax measures currently in place to protect public health from accidental contamination by BSE-contaminated products.

Sunday, September 1, 2013

*** Evaluation of the Zoonotic Potential of Transmissible Mink Encephalopathy

We previously described the biochemical similarities between PrPres derived from L-BSE infected macaque and cortical MM2 sporadic CJD: those observations suggest a link between these two uncommon prion phenotypes in a primate model (it is to note that such a link has not been observed in other models less relevant from the human situation as hamsters or transgenic mice overexpressing ovine PrP [28]). We speculate that a group of related animal prion strains (L-BSE, c-BSE and TME) would have a zoonotic potential and lead to prion diseases in humans with a type 2 PrPres molecular signature (and more specifically type 2B for vCJD)


Together with previous experiments performed in ovinized and bovinized transgenic mice and hamsters [8,9] indicating similarities between TME and L-BSE, the data support the hypothesis that L-BSE could be the origin of the TME outbreaks in North America and Europe during the mid-1900s.




 On June 24, 2005, the USDA announced receipt of final results from The Veterinary Laboratories Agency in Weybridge, England, confirming BSE in a cow that had conflicting test results in 2004. This cow was from Texas, died at approximately 12 years of age, and represented the first endemic case of BSE in the United States. (see Texas BSE Investigation, Final Epidemiology Report, August 2005 External Web Site Policy PDF Document Icon (PDF – 83 KB))




 On March 15, 2006, the USDA announced the confirmation of BSE in a cow in Alabama. The case was identified in a non-ambulatory (downer) cow on a farm in Alabama. The animal was euthanized by a local veterinarian and buried on the farm. The age of the cow was estimated by examination of the dentition as 10-years-old. It had no ear tags or distinctive marks; the herd of origin could not be identified despite an intense investigation (see second featured item above and Alabama BSE Investigation, Final Epidemiology Report, May 2006 External Web Site PolicyPDF Document Icon (PDF – 104 KB)).


 In August 2008, several ARS investigators reported that a rare, genetic abnormality that may persist within the cattle population "is considered to have caused" BSE in this atypical (H-type) BSE animal from Alabama. (See Identification of a Heritable Polymorphism in Bovine PRNP Associated with Genetic Transmissible Spongiform Encephalopathy: Evidence of Heritable BSE External Web Site Policy. Also see BSE Case Associated with Prion Protein Gene Mutation External Web Site Policy.)


 On December 23, 2003, the U.S. Department of Agriculture (USDA) announced a presumptive diagnosis of the first known case of BSE in the United States. It was in an adult Holstein cow from Washington State. This diagnosis was confirmed by an international reference laboratory in Weybridge, England, on December 25. Trace-back based on an ear-tag identification number and subsequent genetic testing confirmed that the BSE-infected cow was imported into the United States from Canada in August 2001. Because the animal was non-ambulatory (a "downer cow") at slaughter, brain tissue samples were taken by USDA's Animal and Plant Health Inspection Service as part of its targeted surveillance for BSE. However the animal's condition was attributed to complications from calving. After the animal was examined by a USDA Food Safety and Inspection Service (FSIS) veterinary medical officer both before and after slaughter, the carcass was released for use as food for human consumption. During slaughter, the tissues considered to be at high risk for the transmission of the BSE agent were removed. On December 24, 2003, FSIS recalled beef from cattle slaughtered in the same plant on the same day as the BSE positive cow. (see Bovine Spongiform Encephalopathy in a Dairy Cow - Washington State, 2003.)



 Tuesday, August 22, 2006







 Unnecessary precautions BSE MAD COW DISEASE Dr. William James FSIS VS Dr. Linda Detwiler 2014



 IF, spontaneous BSE was ever to be proven, it would be the industry, and consumer’s worst nightmare. you could never ever eradicate mad cow disease, no matter how hard you try...terry


 *** Singeltary reply ; Molecular, Biochemical and Genetic Characteristics of BSE in Canada Singeltary reply ;



 LET'S take a closer look at this new prionpathy or prionopathy, and then let's look at the g-h-BSEalabama mad cow. This new prionopathy in humans? the genetic makeup is IDENTICAL to the g-h-BSEalabama mad cow, the only _documented_ mad cow in the world to date like this, ......wait, it get's better. this new prionpathy is killing young and old humans, with LONG DURATION from onset of symptoms to death, and the symptoms are very similar to nvCJD victims, OH, and the plaques are very similar in some cases too, bbbut, it's not related to the g-h-BSEalabama cow, WAIT NOW, it gets even better, the new human prionpathy that they claim is a genetic TSE, has no relation to any gene mutation in that family. daaa, ya think it could be related to that mad cow with the same genetic make-up ??? there were literally tons and tons of banned mad cow protein in Alabama in commerce, and none of it transmitted to cows, and the cows to humans there from ??? r i g h t $$$ ALABAMA MAD COW g-h-BSEalabama In this study, we identified a novel mutation in the bovine prion protein gene (Prnp), called E211K, of a confirmed BSE positive cow from Alabama, United States of America. This mutation is identical to the E200K pathogenic mutation found in humans with a genetic form of CJD. This finding represents the first report of a confirmed case of BSE with a potential pathogenic mutation within the bovine Prnp gene. We hypothesize that the bovine Prnp E211K mutation most likely has caused BSE in "the approximately 10-year-old cow" carrying the E221K mutation.




 Saturday, August 14, 2010


 BSE Case Associated with Prion Protein Gene Mutation (g-h-BSEalabama) and VPSPr PRIONPATHY (see mad cow feed in COMMERCE IN ALABAMA...TSS)



 her healthy calf also carried the mutation


 (J. A. Richt and S. M. Hall PLoS Pathog. 4, e1000156; 2008).


 This raises the possibility that the disease could occasionally be genetic in origin. Indeed, the report of the UK BSE Inquiry in 2000 suggested that the UK epidemic had most likely originated from such a mutation and argued against the scrapierelated assumption. Such rare potential pathogenic PRNP mutations could occur in countries at present considered to be free of BSE, such as Australia and New Zealand. So it is important to maintain strict surveillance for BSE in cattle, with rigorous enforcement of the ruminant feed ban (many countries still feed ruminant proteins to pigs). Removal of specified risk material, such as brain and spinal cord, from cattle at slaughter prevents infected material from entering the human food chain. Routine genetic screening of cattle for PRNP mutations, which is now available, could provide additional data on the risk to the public. Because the point mutation identified in the Alabama animals is identical to that responsible for the commonest type of familial (genetic) CJD in humans, it is possible that the resulting infective prion protein might cross the bovine-human species barrier more easily. Patients with vCJD continue to be identified. The fact that this is happening less often should not lead to relaxation of the controls necessary to prevent future outbreaks.


 Malcolm A. Ferguson-Smith Cambridge University Department of Veterinary Medicine, Madingley Road, Cambridge CB3 0ES, UK e-mail: Jürgen A. Richt College of Veterinary Medicine, Kansas State University, K224B Mosier Hall, Manhattan, Kansas 66506-5601, USA NATURE|Vol 457|26 February 2009









 a) EVSRC Custom dairy feed, Recall # V-130-6;


 b) Performance Chick Starter, Recall # V-131-6;


 c) Performance Quail Grower, Recall # V-132-6;


 d) Performance Pheasant Finisher, Recall # V-133-6.








 Donaldson & Hasenbein/dba J&R Feed Service, Inc., Cullman, AL, by telephone on June 23, 2006 and by letter dated July 19, 2006. Firm initiated recall is complete.




 Dairy and poultry feeds were possibly contaminated with ruminant based protein.




 477.72 tons










 a) Dairy feed, custom, Recall # V-134-6;


 b) Custom Dairy Feed with Monensin, Recall # V-135-6.




 None. Bulk product




 Recalling Firm: Burkmann Feed, Greeneville, TN, by Telephone beginning on June 28, 2006.


 Manufacturer: H. J. Baker & Bro., Inc., Albertville, AL. Firm initiated recall is complete.




 Possible contamination of dairy feeds with ruminant derived meat and bone meal.




 1,484 tons




 TN and WV













 Bulk custom made dairy feed, Recall # V-115-6








 Hiseville Feed & Seed Co., Hiseville, KY, by telephone and letter on or about July 14, 2006. FDA initiated recall is ongoing.




 Custom made feeds contain ingredient called Pro-Lak which may contain ruminant derived meat and bone meal.




 Approximately 2,223 tons










 Bulk custom made dairy feed, Recall # V-116-6








 Rips Farm Center, Tollesboro, KY, by telephone and letter on July 14, 2006. FDA initiated recall is ongoing.




 Custom made feeds contain ingredient called Pro-Lak which may contain ruminant derived meat and bone meal.




 1,220 tons










 Bulk custom made dairy feed, Recall # V-117-6








 Kentwood Co-op, Kentwood, LA, by telephone on June 27, 2006. FDA initiated recall is completed.




 Possible contamination of animal feed ingredients, including ingredients that are used in feed for dairy animals, with ruminant derived meat and bone meal.




 40 tons




 LA and MS






 Bulk Dairy Feed, Recall V-118-6








 Cal Maine Foods, Inc., Edwards, MS, by telephone on June 26, 2006. FDA initiated recall is complete.




 Possible contamination of animal feed ingredients, including ingredients that are used in feed for dairy animals, with ruminant derived meat and bone meal.




 7,150 tons










 Bulk custom dairy pre-mixes, Recall # V-119-6








 Walthall County Co-op, Tylertown, MS, by telephone on June 26, 2006. Firm initiated recall is complete.




 Possible contamination of dairy animal feeds with ruminant derived meat and bone meal.




 87 tons










 Bulk custom dairy pre-mixes, Recall # V-120-6








 Ware Milling Inc., Houston, MS, by telephone on June 23, 2006. Firm initiated recall is complete.




 Possible contamination of dairy animal feeds with ruminant derived meat and bone meal.




 350 tons




 AL and MS






 a) Tucker Milling, LLC Tm 32% Sinking Fish Grower, #2680-Pellet,


 50 lb. bags, Recall # V-121-6;


 b) Tucker Milling, LLC #31120, Game Bird Breeder Pellet,


 50 lb. bags, Recall # V-122-6;


 c) Tucker Milling, LLC #31232 Game Bird Grower,


 50 lb. bags, Recall # V-123-6;


 d) Tucker Milling, LLC 31227-Crumble, Game Bird Starter, BMD Medicated, 50 lb bags, Recall # V-124-6;


 e) Tucker Milling, LLC #31120, Game Bird Breeder, 50 lb bags, Recall # V-125-6;


 f) Tucker Milling, LLC #30230, 30 % Turkey Starter, 50 lb bags, Recall # V-126-6;


 g) Tucker Milling, LLC #30116, TM Broiler Finisher, 50 lb bags, Recall # V-127-6




 All products manufactured from 02/01/2005 until 06/20/2006




 Recalling Firm: Tucker Milling LLC, Guntersville, AL, by telephone and visit on June 20, 2006, and by letter on June 23, 2006.


 Manufacturer: H. J. Baker and Brothers Inc., Stamford, CT. Firm initiated recall is ongoing.




 Poultry and fish feeds which were possibly contaminated with ruminant based protein were not labeled as "Do not feed to ruminants".




 7,541-50 lb bags




 AL, GA, MS, and TN







 Subject: MAD COW FEED RECALL AL AND FL VOLUME OF PRODUCT IN COMMERCE 125 TONS Products manufactured from 02/01/2005 until 06/06/2006


 Date: August 6, 2006 at 6:16 pm PST PRODUCT


 a) CO-OP 32% Sinking Catfish, Recall # V-100-6;


 b) Performance Sheep Pell W/Decox/A/N, medicated, net wt. 50 lbs, Recall # V-101-6;


 c) Pro 40% Swine Conc Meal -- 50 lb, Recall # V-102-6;


 d) CO-OP 32% Sinking Catfish Food Medicated, Recall # V-103-6;


 *** e) "Big Jim's" BBB Deer Ration, Big Buck Blend, Recall # V-104-6;


 f) CO-OP 40% Hog Supplement Medicated Pelleted, Tylosin 100 grams/ton, 50 lb. bag, Recall # V-105-6;


 g) Pig Starter Pell II, 18% W/MCDX Medicated 282020, Carbadox -- 0.0055%, Recall # V-106-6;


 h) CO-OP STARTER-GROWER CRUMBLES, Complete Feed for Chickens from Hatch to 20 Weeks, Medicated, Bacitracin Methylene Disalicylate, 25 and 50 Lbs, Recall # V-107-6;


 i) CO-OP LAYING PELLETS, Complete Feed for Laying Chickens, Recall # 108-6;


 j) CO-OP LAYING CRUMBLES, Recall # V-109-6;


 k) CO-OP QUAIL FLIGHT CONDITIONER MEDICATED, net wt 50 Lbs, Recall # V-110-6;


 l) CO-OP QUAIL STARTER MEDICATED, Net Wt. 50 Lbs, Recall # V-111-6;


 m) CO-OP QUAIL GROWER MEDICATED, 50 Lbs, Recall # V-112-6 CODE


 Product manufactured from 02/01/2005 until 06/06/2006


 RECALLING FIRM/MANUFACTURER Alabama Farmers Cooperative, Inc., Decatur, AL, by telephone, fax, email and visit on June 9, 2006. FDA initiated recall is complete.


 REASON Animal and fish feeds which were possibly contaminated with ruminant based protein not labeled as "Do not feed to ruminants".



















 a) PRO-LAK, bulk weight, Protein Concentrate for Lactating Dairy Animals, Recall # V-079-6;


 b) ProAmino II, FOR PREFRESH AND LACTATING COWS, net weight 50lb (22.6 kg), Recall # V-080-6;




 d) Feather Meal, Recall # V-082-6 CODE


 a) Bulk


 b) None


 c) Bulk


 d) Bulk


 RECALLING FIRM/MANUFACTURER H. J. Baker & Bro., Inc., Albertville, AL, by telephone on June 15, 2006 and by press release on June 16, 2006. Firm initiated recall is ongoing.




 Possible contamination of animal feeds with ruminent derived meat and bone meal.













 Saturday, August 14, 2010


 BSE Case Associated with Prion Protein Gene Mutation (g-h-BSEalabama) and VPSPr PRIONPATHY


 *** (see mad cow feed in COMMERCE IN ALABAMA...TSS)




 Date: September 6, 2006 at 7:58 am PST PRODUCT


 a) EVSRC Custom dairy feed, Recall # V-130-6;


 b) Performance Chick Starter, Recall # V-131-6;


 c) Performance Quail Grower, Recall # V-132-6;


 d) Performance Pheasant Finisher, Recall # V-133-6.


 CODE None RECALLING FIRM/MANUFACTURER Donaldson & Hasenbein/dba J&R Feed Service, Inc., Cullman, AL, by telephone on June 23, 2006 and by letter dated July 19, 2006. Firm initiated recall is complete.




 Dairy and poultry feeds were possibly contaminated with ruminant based protein.









 Saturday, August 14, 2010


 BSE Case Associated with Prion Protein Gene Mutation (g-h-BSEalabama) and VPSPr PRIONPATHY (see mad cow feed in COMMERCE IN ALABAMA...TSS)



 PLOS Singeltary Comment ;


 *** ruminant feed ban for cervids in the United States ? ***


 31 Jan 2015 at 20:14 GMT



 Saturday, January 24, 2015


 Bovine Spongiform Encephalopathy: Atypical Pros and Cons



 Saturday, January 31, 2015


 RAPID ADVICE 17-2014 : Evaluation of the risk for public health of casings in countries with a “negligible risk status for BSE” and on the risk of modification of the list of specified risk materials (SRM) with regard to BSE



 Conclusion/Significance: Our results point to a possibly higher degree of pathogenicity of BASE than classical BSE in primates and also raise a question about a possible link to one uncommon subset of cases of apparently sporadic CJD. Thus, despite the waning epidemic of classical BSE, the occurrence of atypical strains should temper the urge to relax measures currently in place to protect public health from accidental contamination by BSE-contaminated products.








 Perspectives BIOMEDICINE: A Fresh Look at BSE Bruce Chesebro*


 Mad cow disease, or bovine spongiform encephalopathy (BSE), is the cattle form of a family of progressive brain diseases. These diseases include scrapie in sheep, Creutzfeldt-Jakob disease (CJD) in humans, and chronic wasting disease (CWD) in deer and elk. They are also known as either "prion diseases" because of the association of a misfolded cellular prion protein in pathogenesis or "transmissible spongiform encephalopathies" (TSEs) because of the spongelike nature of the damaged brain tissue (1).


 The recent discovery of two BSE-infected cows, one in Canada and one in the United States, has dramatically increased concern in North America among meat producers and consumers alike over the extent to which BSE poses a threat to humans as well as to domestic and wild animals. The European BSE epidemic of the late-1980s seems to have been initiated a decade earlier in the United Kingdom by changes in the production of meat and bone meal (MBM) from rendered livestock, which led to contamination of MBM with the BSE infectious agent. Furthermore, the fact that UK farmers fed this rendered MBM to younger animals and that this MBM was distributed to many countries may have contributed to the ensuing BSE epidemic in the United Kingdom and internationally (2).


 Despite extensive knowledge about the spread of BSE through contaminated MBM, the source of BSE in Europe remains an unsolved mystery (2). It has been proposed that BSE could be derived from a cross-species infection, perhaps through contamination of MBM by scrapie-infected sheep tissues (see the figure). Alternatively, BSE may have been an endemic disease in cattle that went unnoticed because of its low level of horizontal transmission. Lastly, BSE might have originated by "spontaneous" misfolding of the normal cellular prion protein into the disease-associated abnormal isoform (3), which is postulated to be the infectious agent or "prion."


 Five possible sources of BSE in North American cattle. Sheep, deer, and elk could spread prion diseases (TSEs) to cattle through direct animal contact or contamination of pastures. Endemic BSE has not been proven to exist anywhere in the world, but it is difficult to exclude this possibility because of the inefficient spread of BSE infectivity between individual animals (2). BSE caused by spontaneous misfolding of the prion protein has not been proven. CREDIT: KATHARINE SUTLIFF/SCIENCE




 Nevertheless, the idea that BSE might originate due to the spontaneous misfolding of prion proteins has received renewed interest in the wake of reports suggesting the occurrence of atypical BSE (9-11). These results imply that new strains of cattle BSE might have originated separately from the main UK outbreak. Where and how might such strains have originated? Although such rare events cannot be studied directly, any number of sources of the original BSE strain could also explain the discovery of additional BSE strains in cattle (see the figure). However, it would be worrisome if spontaneous BSE were really a valid etiology because such a mechanism would be impossible to prevent--unlike other possible scenarios that could be controlled by large-scale eradication of TSE-positive animals.


 Another way to look at this problem is to examine evidence for possible spontaneous TSE disease in other animals besides cattle. Spontaneous BSE would be extremely difficult to detect in cattle, where horizontal spread is minimal. However, in the case of the sheep TSE disease, scrapie, which spreads from ewes to lambs at birth as well as between adults, spontaneous disease should be detectable as new foci of clinical infection. In the early 1950s scrapie was eradicated in both Australia and New Zealand, and the mainland of both these countries has remained scrapie-free ever since. This scrapie-free status is not the result of selection of sheep resistant to scrapie because sheep from New Zealand are as susceptible as their UK counterparts to experimental scrapie infection (12). These experiments of man and nature appear to indicate that spontaneous clinical scrapie does not occur in sheep. Similarly, because CWD is known to spread horizontally, the lack of CWD in the deer or elk of eastern North America but its presence in western regions would also argue against a spontaneous disease mechanism. This is particularly noteworthy in New Zealand, where there are large numbers of deer and elk farms and yet no evidence of spontaneous CWD. If spontaneous scrapie does not occur in sheep or deer, this would suggest that spontaneous forms of BSE and sporadic Creutzfeldt-Jakob disease (sCJD) are unlikely to be found in cattle or humans. The main caveat to this notion is that spontaneous disease may arise in some animal species but not others. In humans, sCJD--which is considered by some researchers to begin by spontaneous misfolding of the prion protein--usually takes more than 50 years to appear. Thus, in animals with a shorter life-span, such as sheep, deer, and cattle, an analogous disease mechanism might not have time to develop.


 What can we conclude so far about BSE in North America? Is the BSE detected in two North American cows sporadic or spontaneous or both? "Sporadic" pertains to the rarity of disease occurrence. "Spontaneous" pertains to a possible mechanism of origin of the disease. These are not equivalent terms. The rarity of BSE in North America qualifies it as a sporadic disease, but this low incidence does not provide information about cause. For the two reported North American BSE cases, exposure to contaminated MBM remains the most likely culprit. However, other mechanisms are still possible, including cross-infection by sheep with scrapie or cervids with CWD, horizontal transmission from cattle with endemic BSE, and spontaneous disease in individual cattle. Based on our understanding of other TSEs, the spontaneous mechanism is probably the least likely. Thus, "idiopathic" BSE--that is, BSE of unknown etiology--might be a better term to describe the origin of this malady. ...


 snip...full text ;



 DR. DEHAVEN: “All right. I think we've got three different questions in there, and I'll try to touch on each one of them.


 “First of all, let me correct just a technical issue, and that is you mentioned 1 in 10,000. And actually our surveillance system currently is designed, the one that we have in place now is designed to detect 1 positive in 1 million cattle, and I gave some numbers between 200,000 and 268,000 that would allow us to detect 1 in 10 million as opposed to 1 in 10,000.


 “So we would, if we were able to collect in the ballpark of those numbers of samples then we with increasing numbers of samples have an increasingly statistically valid sample from which to determine, one, whether or not the disease exists and, if so, at what prevalence level.


 “So our real emphasis is to test as many of those animals as we can, ensure that we get an appropriate geographical distribution, but not setting a specific number as far as a target. Again, consistent with the recommendation from the International Review Team, their recommendation was to test all of them.


 “So that's consistent with where we're going is to test as many as we possibly can.


 *** “As far as spontaneous cases, that is a very difficult issue. There is no evidence to prove that spontaneous BSE occurs in cattle; but here again it's an issue of proving a negative. We do know that CJD, the human version of the disease, does occur spontaneously in humans at the rate of about 1 in 1 million. We don't have enough data to definitively say that spontaneous cases of BSE in cattle occur or do not occur.


 “Again, it's a very difficult situation to prove a negative.


 “So a lot of research is ongoing. Certainly if we do come up with any positive samples in the course of this surveillance we will be looking at that question in evaluating those samples but no scientifically hard evidence to confirm or refute whether or not spontaneous cases of BSE occur.






 1. The BSE epidemic


 1.1. The origin of the BSE epidemic will probably never be determined with certainty.


 1.2. We do not know whether or not some of the BARB cases represent truly sporadic classical BSE. If there are spontaneous cases then BSE will never be eradicated although reducing surveillance could make it appear that BSE has been eradicated.




 5.3. It was stated that the number of sporadic CJD cases was rising. Participants were invited to discuss the reason for this. It was suggested that this was likely to be due to improved surveillance with more cases of sporadic CJD being detected (i.e. through MRI scans). There had been a similar increase in sporadic CJD in countries which did not have a BSE epidemic but improved their surveillance. This supported this theory and suggested that the increase in sporadic CJD was not related to the BSE outbreak.



 Atypical BSE: Transmissibility


 Linda Detwiller, 5/10/2011


  BASE (L) transmitted to:  cattle (IC) - inc < 20 mos and oral?)


  Cynomolgus macaques (IC)


  Mouse lemurs (IC and oral)


  wild-type mice (IC)


  bovinized transgenic mice (IC and IP)


  humanized transgenic mice (IC)


  H cases transmitted to:


  cattle – IC incubations < 20 months


  bovinized transgenic mice (IC)


  ovinized transgenic mice (IC)


  C57BL mice (IC)


  One study did not transmit to humanized PrP Met 129 mice


 Evaluation of Possibility of Atypical


 BSE Transmitting to Humans


  Possble interpretation:


  L type seems to transmit to nonhuman primates with greater ease than classical BSE


  L type also transmitted to humanized transgenic mice with higher attack rate and shorter incubation period than classical?


  H type did not transmit to Tg Hu transgenic mice


 Linda Detwiller, 5/10/2011



 I ask Professor Kong ;


 Thursday, December 04, 2008 3:37 PM


 Subject: RE: re--Chronic Wating Disease (CWD) and Bovine Spongiform Encephalopathies (BSE): Public Health Risk Assessment


 IS the h-BSE more virulent than typical BSE as well, or the same as cBSE, or less virulent than cBSE? just curious.....


 Professor Kong reply ;




 As to the H-BSE, we do not have sufficient data to say one way or another, but we have found that H-BSE can infect humans. I hope we could publish these data once the study is complete. Thanks for your interest.


 Best regards, Qingzhong Kong, PhD Associate Professor Department of Pathology Case Western Reserve University Cleveland, OH 44106 USA


 BSE-H is also transmissible in our humanized Tg mice. The possibility of more than two atypical BSE strains will be discussed.


 Supported by NINDS NS052319, NIA AG14359, and NIH AI 77774.




 P.4.23 Transmission of atypical BSE in humanized mouse models


 Liuting Qing1, Wenquan Zou1, Cristina Casalone2, Martin Groschup3, Miroslaw Polak4, Maria Caramelli2, Pierluigi Gambetti1, Juergen Richt5, Qingzhong Kong1 1Case Western Reserve University, USA; 2Instituto Zooprofilattico Sperimentale, Italy; 3Friedrich-Loeffler-Institut, Germany; 4National Veterinary Research Institute, Poland; 5Kansas State University (Previously at USDA National Animal Disease Center), USA


 Background: Classical BSE is a world-wide prion disease in cattle, and the classical BSE strain (BSE-C) has led to over 200 cases of clinical human infection (variant CJD). Atypical BSE cases have been discovered in three continents since 2004; they include the L-type (also named BASE), the H-type, and the first reported case of naturally occurring BSE with mutated bovine PRNP (termed BSE-M). The public health risks posed by atypical BSE were argely undefined.


 Objectives: To investigate these atypical BSE types in terms of their transmissibility and phenotypes in humanized mice.


 Methods: Transgenic mice expressing human PrP were inoculated with several classical (C-type) and atypical (L-, H-, or Mtype) BSE isolates, and the transmission rate, incubation time, characteristics and distribution of PrPSc, symptoms, and histopathology were or will be examined and compared.


 Results: Sixty percent of BASE-inoculated humanized mice became infected with minimal spongiosis and an average incubation time of 20-22 months, whereas only one of the C-type BSE-inoculated mice developed prion disease after more than 2 years. Protease-resistant PrPSc in BASE-infected humanized Tg mouse brains was biochemically different from bovine BASE or sCJD. PrPSc was also detected in the spleen of 22% of BASE-infected humanized mice, but not in those infected with sCJD. Secondary transmission of BASE in the humanized mice led to a small reduction in incubation time. The atypical BSE-H strain is also transmissible with distinct phenotypes in the humanized mice, but no BSE-M transmission has been observed so far.


 Discussion: Our results demonstrate that BASE is more virulent than classical BSE, has a lymphotropic phenotype, and displays a modest transmission barrier in our humanized mice. BSE-H is also transmissible in our humanized Tg mice. The possibility of more than two atypical BSE strains will be discussed.


 Supported by NINDS NS052319, NIA AG14359, and NIH AI 77774.





 14th International Congress on Infectious Diseases H-type and L-type Atypical BSE January 2010 (special pre-congress edition)


 18.173 page 189


 Experimental Challenge of Cattle with H-type and L-type Atypical BSE


 A. Buschmann1, U. Ziegler1, M. Keller1, R. Rogers2, B. Hills3, M.H. Groschup1. 1Friedrich-Loeffler-Institut, Greifswald-Insel Riems, Germany, 2Health Canada, Bureau of Microbial Hazards, Health Products & Food Branch, Ottawa, Canada, 3Health Canada, Transmissible Spongiform Encephalopathy Secretariat, Ottawa, Canada


 Background: After the detection of two novel BSE forms designated H-type and L-type atypical BSE the question of the pathogenesis and the agent distribution of these two types in cattle was fully open. From initial studies of the brain pathology, it was already known that the anatomical distribution of L-type BSE differs from that of the classical type where the obex region in the brainstem always displays the highest PrPSc concentrations. In contrast in L-type BSE cases, the thalamus and frontal cortex regions showed the highest levels of the pathological prion protein, while the obex region was only weakly involved.


 Methods:We performed intracranial inoculations of cattle (five and six per group) using 10%brainstemhomogenates of the two German H- and L-type atypical BSE isolates. The animals were inoculated under narcosis and then kept in a free-ranging stable under appropriate biosafety conditions. At least one animal per group was killed and sectioned in the preclinical stage and the remaining animals were kept until they developed clinical symptoms. The animals were examined for behavioural changes every four weeks throughout the experiment following a protocol that had been established during earlier BSE pathogenesis studies with classical BSE.


 Results and Discussion: All animals of both groups developed clinical symptoms and had to be euthanized within 16 months. The clinical picture differed from that of classical BSE, as the earliest signs of illness were loss of body weight and depression. However, the animals later developed hind limb ataxia and hyperesthesia predominantly and the head. Analysis of brain samples from these animals confirmed the BSE infection and the atypical Western blot profile was maintained in all animals. Samples from these animals are now being examined in order to be able to describe the pathoge esis and agent distribution for these novel BSE types.


 Conclusions: A pilot study using a commercially avaialble BSE rapid test ELISA revealed an essential restriction of PrPSc to the central nervous system for both atypical BSE forms. A much more detailed analysis for PrPSc and infectivity is still ongoing.



 14th ICID International Scientific Exchange Brochure - Final Abstract Number: ISE.114


 Session: International Scientific Exchange


 Transmissible Spongiform encephalopathy (TSE) animal and human TSE in North America update October 2009


 T. Singeltary Bacliff, TX, USA


 Background: An update on atypical BSE and other TSE in North America. Please remember, the typical U.K. c-BSE, the atypical l-BSE (BASE), and h-BSE have all been documented in North America, along with the typical scrapie's, and atypical Nor-98 Scrapie, and to date, 2 different strains of CWD, and also TME. All these TSE in different species have been rendered and fed to food producing animals for humans and animals in North America (TSE in cats and dogs ?), and that the trading of these TSEs via animals and products via the USA and Canada has been immense over the years, decades.


 Methods: 12 years independent research of available data


 Results: I propose that the current diagnostic criteria for human TSEs only enhances and helps the spreading of human TSE from the continued belief of the UKBSEnvCJD only theory in 2009. With all the science to date refuting it, to continue to validate this old myth, will only spread this TSE agent through a multitude of potential routes and sources i.e. consumption, medical i.e., surgical, blood, dental, endoscopy, optical, nutritional supplements, cosmetics etc.


 Conclusion: I would like to submit a review of past CJD surveillance in the USA, and the urgent need to make all human TSE in the USA a reportable disease, in every state, of every age group, and to make this mandatory immediately without further delay. The ramifications of not doing so will only allow this agent to spread further in the medical, dental, surgical arena's. Restricting the reporting of CJD and or any human TSE is NOT scientific. Iatrogenic CJD knows NO age group, TSE knows no boundaries. I propose as with Aguzzi, Asante, Collinge, Caughey, Deslys, Dormont, Gibbs, Gajdusek, Ironside, Manuelidis, Marsh, et al and many more, that the world of TSE Transmissible Spongiform Encephalopathy is far from an exact science, but there is enough proven science to date that this myth should be put to rest once and for all, and that we move forward with a new classification for human and animal TSE that would properly identify the infected species, the source species, and then the route.



 Diagnosis and Reporting of Creutzfeldt-Jakob Disease


 Singeltary, Sr et al. JAMA.2001; 285: 733-734. Vol. 285 No. 6, February 14, 2001 JAMA


 Diagnosis and Reporting of Creutzfeldt-Jakob Disease


 To the Editor: In their Research Letter, Dr Gibbons and colleagues1 reported that the annual US death rate due to Creutzfeldt-Jakob disease (CJD) has been stable since 1985. These estimates, however, are based only on reported cases, and do not include misdiagnosed or preclinical cases. It seems to me that misdiagnosis alone would drastically change these figures. An unknown number of persons with a diagnosis of Alzheimer disease in fact may have CJD, although only a small number of these patients receive the postmortem examination necessary to make this diagnosis. Furthermore, only a few states have made CJD reportable. Human and animal transmissible spongiform encephalopathies should be reportable nationwide and internationally.


 Terry S. Singeltary, Sr Bacliff, Tex


 1. Gibbons RV, Holman RC, Belay ED, Schonberger LB. Creutzfeldt-Jakob disease in the United States: 1979-1998. JAMA. 2000;284:2322-2323.



 26 March 2003


 Terry S. Singeltary, retired (medically) CJD WATCH


 I lost my mother to hvCJD (Heidenhain Variant CJD). I would like to comment on the CDC's attempts to monitor the occurrence of emerging forms of CJD. Asante, Collinge et al [1] have reported that BSE transmission to the 129-methionine genotype can lead to an alternate phenotype that is indistinguishable from type 2 PrPSc, the commonest sporadic CJD. However, CJD and all human TSEs are not reportable nationally. CJD and all human TSEs must be made reportable in every state and internationally. I hope that the CDC does not continue to expect us to still believe that the 85%+ of all CJD cases which are sporadic are all spontaneous, without route/source. We have many TSEs in the USA in both animal and man. CWD in deer/elk is spreading rapidly and CWD does transmit to mink, ferret, cattle, and squirrel monkey by intracerebral inoculation. With the known incubation periods in other TSEs, oral transmission studies of CWD may take much longer. Every victim/family of CJD/TSEs should be asked about route and source of this agent. To prolong this will only spread the agent and needlessly expose others. In light of the findings of Asante and Collinge et al, there should be drastic measures to safeguard the medical and surgical arena from sporadic CJDs and all human TSEs. I only ponder how many sporadic CJDs in the USA are type 2 PrPSc?



 2 January 2000


 British Medical Journal


 U.S. Scientist should be concerned with a CJD epidemic in the U.S., as well



 15 November 1999


 British Medical Journal


 vCJD in the USA * BSE in U.S.



 The Lancet Infectious Diseases, Volume 3, Issue 8, Page 463, August 2003 doi:10.1016/S1473-3099(03)00715-1Cite or Link Using DOI


 Tracking spongiform encephalopathies in North America




 Xavier Bosch


 “My name is Terry S Singeltary Sr, and I live in Bacliff, Texas. I lost my mom to hvCJD (Heidenhain variant CJD) and have been searching for answers ever since. What I have found is that we have not been told the truth. CWD in deer and elk is a small portion of a much bigger problem.” 49-year—old Singeltary is one of a number of people who have remained largely unsatisfied after being told that a close relative died from a rapidly progressive dementia compatible with spontaneous Creutzfeldt—Jakob ...



 Suspect symptoms


 What if you can catch old-fashioned CJD by eating meat from a sheep infected with scrapie?


 28 Mar 01 Most doctors believe that sCJD is caused by a prion protein deforming by chance into a killer. But Singeltary thinks otherwise. He is one of a number of campaigners who say that some sCJD, like the variant CJD related to BSE, is caused by eating meat from infected animals. Their suspicions have focused on sheep carrying scrapie, a BSE-like disease that is widespread in flocks across Europe and North America.


 Now scientists in France have stumbled across new evidence that adds weight to the campaigners' fears. To their complete surprise, the researchers found that one strain of scrapie causes the same brain damage in mice as sCJD.


 "This means we cannot rule out that at least some sCJD may be caused by some strains of scrapie," says team member Jean-Philippe Deslys of the French Atomic Energy Commission's medical research laboratory in Fontenay-aux-Roses, south-west of Paris. Hans Kretschmar of the University of Göttingen, who coordinates CJD surveillance in Germany, is so concerned by the findings that he now wants to trawl back through past sCJD cases to see if any might have been caused by eating infected mutton or lamb...



 Self-Propagative Replication of Ab Oligomers Suggests Potential Transmissibility in Alzheimer Disease


 Received July 24, 2014; Accepted September 16, 2014; Published November 3, 2014



 Singeltary comment ;



 Saturday, August 14, 2010


 ***BSE Case Associated with Prion Protein Gene Mutation (g-h-BSEalabama) and VPSPr PRIONPATHY


 *** (see mad cow feed in COMMERCE IN ALABAMA...TSS)



 *** What irks many scientists is the USDA’s April 25 statement that the rare disease is “not generally associated with an animal consuming infected feed.”


 The USDA’s conclusion is a “gross oversimplification,” said Dr. Paul Brown, one of the world’s experts on this type of disease who retired recently from the National Institutes of Health. "(The agency) has no foundation on which to base that statement.”



 Owens, Julie


 From: Terry S. Singeltary Sr. []


 Sent: Monday, July 24, 2006 1:09 PM


 To: FSIS RegulationsComments


 Subject: [Docket No. FSIS-2006-0011] FSIS Harvard Risk Assessment of Bovine Spongiform Encephalopathy (BSE) Page 1 of 98






 FDA et al thinks as much as 5.5 grams of SRM is just fine for a heifer weighing about 600 lbs ;


 ''FDA has determined that each animal could have consumed, at most and in total, five-and-one-half grams - approximately a quarter ounce -- of prohibited material. These animals weigh approximately 600 pounds.''



 Saturday, December 21, 2013


 **** Complementary studies detecting classical bovine spongiform encephalopathy infectivity in jejunum, ileum and ileocaecal junction in incubating cattle ****



 Wednesday, May 2, 2012





 Saturday, June 12, 2010


 PUBLICATION REQUEST AND FOIA REQUEST Project Number: 3625-32000-086-05 Study of Atypical Bse



 Wednesday, July 28, 2010


 re-Freedom of Information Act Project Number 3625-32000-086-05, Study of Atypical BSE UPDATE July 28, 2010







 Spread of BSE prions in cynomolgus monkeys (Macaca fascicularis) after oral transmission


 Edgar Holznagel1, Walter Schulz-Schaeffer2, Barbara Yutzy1, Gerhard Hunsmann3, Johannes Loewer1 1Paul-Ehrlich-Institut, Federal Institute for Sera and Vaccines, Germany; 2Department of Neuropathology, Georg-August University, Göttingen, Germany, 3Department of Virology and Immunology, German Primate Centre, Göttingen, Germany


 Background: BSE-infected cynomolgus monkeys represent a relevant animal model to study the pathogenesis of variant Creutzfeldt-Jacob disease (vCJD).


 Objectives: To study the spread of BSE prions during the asymptomatic phase of infection in a simian animal model.


 Methods: Orally BSE-dosed macaques (n=10) were sacrificed at defined time points during the incubation period and 7 orally BSE-dosed macaques were sacrificed after the onset of clinical signs. Neuronal and non-neuronal tissues were tested for the presence of proteinase-K-resistant prion protein (PrPres) by western immunoblot and by paraffin-embedded tissue (PET) blot technique.


 Results: In clinically diseased macaques (5 years p.i. + 6 mo.), PrPres deposits were widely spread in neuronal tissues (including the peripheral sympathetic and parasympathetic nervous system) and in lymphoid tissues including tonsils. In asymptomatic disease carriers, PrPres deposits could be detected in intestinal lymph nodes as early as 1 year p.i., but CNS tissues were negative until 3 – 4 years p.i. Lumbal/sacral segments of the spinal cord and medulla oblongata were PrPres positive as early as 4.1 years p.i., whereas sympathetic trunk and all thoracic/cervical segments of the spinal cord were still negative for PrPres. However, tonsil samples were negative in all asymptomatic cases.


 Discussion: There is evidence for an early spread of BSE to the CNS via autonomic fibres of the splanchnic and vagus nerves indicating that trans-synaptical spread may be a time-limiting factor for neuroinvasion. Tonsils were predominantly negative during the main part of the incubation period indicating that epidemiological vCJD screening results based on the detection of PrPres in tonsil biopsies may mostly tend to underestimate the prevalence of vCJD among humans.



 Simian vCJD can be easily triggered in cynomolgus monkeys on the oral route using less than 5 g BSE brain homogenate.



 Atypical L-Type Bovine Spongiform Encephalopathy (L-BSE) Transmission to Cynomolgus Macaques, a Non-Human Primate


 Jpn. J. Infect. Dis., 64 (1), 81-84, 2011


 To see a printable version of the article in the Adobe file format, click this [PDF] link.


 Short Communication


 Atypical L-Type Bovine Spongiform Encephalopathy (L-BSE) Transmission to Cynomolgus Macaques, a Non-Human Primate


 Fumiko Ono, Naomi Tase1, Asuka Kurosawa3, Akio Hiyaoka, Atsushi Ohyama, Yukio Tezuka, Naomi Wada2, Yuko Sato3, Minoru Tobiume3, Ken'ichi Hagiwara4, Yoshio Yamakawa4*, Keiji Terao1, and Tetsutaro Sata3


 The Corporation for Production and Research of Laboratory Primates, Tsukuba 305-0843; 1Tsukuba Primate Research Center, National Institute of Biomedical Innovation, Tsukuba 305-0843; 2Department of Veterinary Medicine, Yamaguchi University, Yamaguchi 753-8515; and 3Department of Pathology and 4Department of Biochemistry and Cell Biology, National Institute of Infectious Diseases, Tokyo 162-8640, Japan


 (Received December 9, 2010. Accepted December 22, 2010)




 *Corresponding author: Mailing address: Department of Biochemistry and Cell Biology, National Institute of Infectious Diseases, Toyama 1-23-1, Shinjuku-ku, Tokyo 162-8640, Japan. Tel: +81-3-5285-1111 ext. 2127, Fax: +81-3-5285-1157, E-mail:




 SUMMARY: A low molecular weight type of atypical bovine spongiform encephalopathy (L-BSE) was transmitted to two cynomolgus macaques by intracerebral inoculation of a brain homogenate of cattle with atypical BSE detected in Japan. They developed neurological signs and symptoms at 19 or 20 months post-inoculation and were euthanized 6 months after the onset of total paralysis. Both the incubation period and duration of the disease were shorter than those for experimental transmission of classical BSE (C-BSE) into macaques. Although the clinical manifestations, such as tremor, myoclonic jerking, and paralysis, were similar to those induced upon C-BSE transmission, no premonitory symptoms, such as hyperekplexia and depression, were evident. Most of the abnormal prion protein (PrPSc) was confined to the tissues of the central nervous system, as determined by immunohistochemistry and Western blotting. The PrPSc glycoform that accumulated in the monkey brain showed a similar profile to that of L-BSE and consistent with that in the cattle brain used as the inoculant. PrPSc staining in the cerebral cortex showed a diffuse synaptic pattern by immunohistochemistry, whereas it accumulated as fine and coarse granules and/or small plaques in the cerebellar cortex and brain stem. Severe spongiosis spread widely in the cerebral cortex, whereas florid plaques, a hallmark of variant Creutzfeldt-Jakob disease in humans, were observed in macaques inoculated with C-BSE but not in those inoculated with L-BSE.




 To date, 27 cases of L-BSE and 24 cases of H-BSE have been report­ed worldwide (16), thus meaning that the prevalence of atypical BSE is considerably lower than that of C-BSE. However, recent studies showed that L-BSE is easily transmissible to transgenic mice expressing human (17,18) or bovine (19,20) prion protein, as well as to non-human primates (21), with shorter incubation periods than for the transmission of C-BSE to these animals. The virulent nature of L-BSE has stimulated new concern for human public health since the transmis­sion of C-BSE to humans resulted in variant Creutz­feldt-Jakob disease (vCJD) (4-7), a new emergent prion disease.




 Two macaques simultaneously developed neurologi­cal signs and symptoms 19-20 months post-inoculation (mpi) with the brain homogenate of BSEI JP24. The monkeys entered the terminal stage of the disease (total paralysis) at 24-25 mpi, Both the onset and duration of the disease were shorter than those reported for the transmission of C-BSE to macaques by us and other groups (27,28). The clinical manifestations such as tremor, myoclonic jerking, and paralysis were similar to those observed during the transmission of C-BSE to ma- caques, whereas the premonitory abnormal behaviors, such as hyperekplexia and depression, seen upon trans­mission of C-BSE to macaques were not evident (27).


 Histopathological analysis and IHC, performed as described previously (29), showed that severe spon­giform changes and the accumulation of Prpsc with various patterns were detectable in the brains of both monkeys (Fig. 1). Vacuolization was profound throughout the cerebral cortex, from the frontal to the occipital lobes (Fig. la). Likewise, synaptic-type Prpsc precipitation (30) was observed in the whole cerebral cortex and basal ganglia by IHC (Figs. Ib and c). Dense precipitates and plaques of Prpsc, which had been ob­served in cattle (JP24) brain (13), were not detected in the cerebrum of the monkeys. Prpsc, in the form of small plaques or coarse granules, was, however, detect­ed in the molecular layer of the cerebellum (Fig. Ie). Despite the severe spongiosis in the cerebral cortex, florid plaques, which are large Prpsc plaques surround­ed by vacuoles, a hallmark ofvCJD (4-7,30) and C-BSE transmission to macaques (27,28), were not observed. The histopathology of the brain was therefore similar to that reported for the brain of L-BSE (BASE)-transmit­ted macaques (21).




 see full text ;



 Monday, September 13, 2010


 atypical BSE strains and sporadic CJD strains, is there a connection and why shouldn't there be $



 Monday, September 13, 2010


 atypical BSE strains and sporadic CJD strains, is there a connection and why shouldn't there be $ A Surprisingly High Number of the Plaque-Like VV sCJD Subtype Among the Polish sCJD-is There a Connection with BASE?




 A Surprisingly High Number of the Plaque-Like VV sCJD Subtype Among the Polish sCJD—is There a Connection with BASE?


 Beata Sikorska and Pawel P. Liberski Department of Molecular Pathology and Neuropathology; Medical University of Lodz; Lodz, Poland


 Recently described bovine amyloidotic spongiform encephalopathy (BASE) or L type BSE—was is overrepresented in Poland (15% of all cases of BSE). Moreover, the number of BASE cases in Poland per million bovines is the highest in Europe. A potential human risk from BASE is evident from experimental transmission to “humanized” transgenic animals and primates. Taking into consideration that non-human primate inoculated with BASE had a shorter incubation period than monkeys infected with classical BSE, and that humanized Tg mice have been found to be highly susceptible to infection with atypical form of BSE, it seems probable that BASE may be more pathogenic for humans than BSE, but the transmitted disease may differ from BSE-derived vCJD. Among 47 cases which have been diagnosed as definite in our laboratory, in 19 cases complete histopathological examination and codon 129 status were available. On the basis of the histological pattern and codon 129 status the cases of sCJD were divided into subtypes according to the Parchi&Gambetti classification. The results are as follows: type 1 (MMorMV)- 42%, type 2 (VV)-32%, type 3 (MV)-10.5%, type 4c (MM)- 10.5% and type 5 (VV)-5 %. Although the number of cases is too low to conclude a significantly different distribution of sCJD subtypes in Polish population those data show surprisingly high number of the plaque-like VV sCJD subtype. Interestingly, it was shown before that Tg mice inoculated with BASE showed granular and plaque-like aggregates or PrPSc in brains resembling those observed in VV2 subtype of sCJD.




 Transmission of Classical and Atypical (L-type) Bovine Spongiform Encephalopathy (BSE) Prions to Cynomolgus macaques


 Fumiko Ono,1 Yoshio Yamakawa,2 Minoru Tobiume,3 Yuko Sato,3 Harutaka Katano,3 Kenichi Hagiwara,2 Iori Itagaki,1 Akio Hiyaoka,1 Katuhiko Komatuzaki,1 Yasunori Emoto,1 Hiroaki Shibata,4 Yuichi Murayama,5 Keiji Terao,4 Yasuhiro Yasutomi4 and Tetsutaro Sata3


 1The Corporation for Production and Research of Laboratory Primates; Tsukuba City, Japan; 2Departments of Cell Biology and Biochemistry; and 3Pathology; National Institute of Infectious Diseases; Tokyo, Japan; 4Tsukuba Primate Research Center; National Institute of Biomedical Innovation; Tsukuba City, Japan; 5Prion Disease Research Team; National Institute of Animal Health; Tsukuba City, Japan


 Key words: L-type BSE, cBSE, cynomolgus macaques, transmission


 BSE prion derived from classical BSE (cBSE) or L-type BSE was characterized by inoculation into the brain of cynomolgus macaques. The neurologic manifestation was developed in all cynomolgus macaques at 27–43 months after intracerebral inoculation of brain homogenate from cBSE-affected cattle (BSE JP/6). Second transmission of cBSE from macaque to macaque shortened incubation period to 13–18 months. cBSE-affected macaques showed the similar clinical signs including hyperekplexia, tremor and paralysis in both primary and second transmission.


 Two macaques were intracerebrally inoculated brain homogenate from the L-type BSE-affected cattle (BSE JP/24). The incubation periods were 19–20 months in primary transmission.


 The clinical course of the L-type BSE-affected macaques differed from that in cBSE-affected macaques in the points of severe myoclonus without hyperekplexia. The glycoform profile of PrPSc detected in macaque CNS was consistent with original pattern of either cBSE or L-typeBSE PrPSc, respectively. Although severe spongiform change in the brain was remarkable in all BSE-affected macaques, severe spongiform spread widely in cerebral cortex in L-type BSE-affected macaques. Heavy accumulation of PrPSc surrounded by vacuola formed florid plaques in cerebral cortex of cBSE-affected macaques. Deposit of PrPSc in L-type BSE-affected macaque was weak and diffuse synaptic pattern in cerebrum, but large PrPSc plaques were evident at cerebellum. MRI analysis, T2, T1, DW and flair sequences, at the time of autopsy revealed that brain atrophy and dilatation of cerebral ventricles were significantly severe in L-type BSE-affected macaques. These results suggest that L-type BSE is more virulent strain to primates comparing to cBSE.




 Evidence from Molecular Strain Typing


 Gianluigi Zanusso Department of Neurological and Visual Sciences; Section of Clinical Neurology; University of Verona; Verona, Italy


 Key words: molecular analysis, strain typing, atypical BSE, CJD


 In 2001, active surveillance for bovine spongiform encephalopathy (BSE) led to the discovery of atypical BSE phenotypes in aged cattle distinct from classical BSE (C-type). These atypical BSE cases had been classified as low L-type (BASE) or high H-type BSE based on the molecular mass and the degree of glycosylation of of the pathological prion protein (PrPSc). Transmission studies in TgBov mice showed that H-type BSE, C-type BSE and BASE behave as distinct prion strains with different incubation periods, PrPSc molecular patterns and pathological phenotypes. A still unclear issue concerns the potential transmissibility and phenotypes of atypical BSEs in humans. We previously indicated that BASE was similar to a distinct subgroup of sporadic form of Creutzfeldt-Jakob disease (sCJD) MV2, based on molecular similarities and on neuropathological pattern of PrP deposition. To investigate a possible link between BASE and sCJD, Kong et al. and Comoy et al. experimentally inoculated TgHu mice (129MM) and a non-human primate respectively, showing in both models that BASE was more virulent compare to BSE. Further, non-human primate reproduced a clinical phenotype resembling to that of sCJD subtype MM2. Here, we presented a comparative analysis of the biochemical fingerprints of PrPSc between the different sCJD subtypes and animal TSEs and after experimental transmission to animals.



 Opinion of the Scientific Steering Committee on the GEOGRAPHICAL RISK OF BOVINE SPONGIFORM ENCEPHALOPATHY (GBR) in POLAND Adopted on 30/03/2001


 It is concluded that it is likely but not confirmed that one or several cattle that are (pre-clinically or clinically) infected with the BSE agent are currently present in the domestic herd of Poland (GBR III).




 Atypical BSE (BASE) Transmitted from Asymptomatic Aging Cattle to a Primate


 Emmanuel E. Comoy1*, Cristina Casalone2, Nathalie Lescoutra-Etchegaray1, Gianluigi Zanusso3, Sophie Freire1, Dominique Marcé1, Frédéric Auvré1, Marie-Magdeleine Ruchoux1, Sergio Ferrari3, Salvatore Monaco3, Nicole Salès4, Maria Caramelli2, Philippe Leboulch1,5, Paul Brown1, Corinne I. Lasmézas4, Jean-Philippe Deslys1


 1 Institute of Emerging Diseases and Innovative Therapies, CEA, Fontenay-aux-Roses, France, 2 Istituto Zooprofilattico Sperimentale del Piemonte, Turin, Italy, 3 Policlinico G.B. Rossi, Verona, Italy, 4 Scripps Florida, Jupiter, Florida, United States of America, 5 Genetics Division, Brigham & Women's Hospital, Harvard Medical School, Boston, Massachusetts, United States of America


 Abstract Top Background Human variant Creutzfeldt-Jakob Disease (vCJD) results from foodborne transmission of prions from slaughtered cattle with classical Bovine Spongiform Encephalopathy (cBSE). Atypical forms of BSE, which remain mostly asymptomatic in aging cattle, were recently identified at slaughterhouses throughout Europe and North America, raising a question about human susceptibility to these new prion strains.


 Methodology/Principal Findings Brain homogenates from cattle with classical BSE and atypical (BASE) infections were inoculated intracerebrally into cynomolgus monkeys (Macacca fascicularis), a non-human primate model previously demonstrated to be susceptible to the original strain of cBSE. The resulting diseases were compared in terms of clinical signs, histology and biochemistry of the abnormal prion protein (PrPres). The single monkey infected with BASE had a shorter survival, and a different clinical evolution, histopathology, and prion protein (PrPres) pattern than was observed for either classical BSE or vCJD-inoculated animals. Also, the biochemical signature of PrPres in the BASE-inoculated animal was found to have a higher proteinase K sensitivity of the octa-repeat region. We found the same biochemical signature in three of four human patients with sporadic CJD and an MM type 2 PrP genotype who lived in the same country as the infected bovine.


 Conclusion/Significance Our results point to a possibly higher degree of pathogenicity of BASE than classical BSE in primates and also raise a question about a possible link to one uncommon subset of cases of apparently sporadic CJD. Thus, despite the waning epidemic of classical BSE, the occurrence of atypical strains should temper the urge to relax measures currently in place to protect public health from accidental contamination by BSE-contaminated products.


 Citation: Comoy EE, Casalone C, Lescoutra-Etchegaray N, Zanusso G, Freire S, et al. (2008) Atypical BSE (BASE) Transmitted from Asymptomatic Aging Cattle to a Primate. PLoS ONE 3(8): e3017. doi:10.1371/journal.pone.0003017


 Editor: Neil Mabbott, University of Edinburgh, United Kingdom


 Received: April 24, 2008; Accepted: August 1, 2008; Published: August 20, 2008


 Copyright: © 2008 Comoy et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.


 Funding: This work has been supported by the Network of Excellence NeuroPrion.


 Competing interests: CEA owns a patent covering the BSE diagnostic tests commercialized by the company Bio-Rad.


 * E-mail:



 Session I - Prions: Structure, Strain and Detection (II)


 Searching for BASE Strain Signature in Sporadic Creutzfedlt-Jakob Disease


 Gianluigi Zanusso


 Department of Neurological and Visual Sciences, Section of Clinical Neurology University of Verona, Verona, Italy.


 Bovine amyloidotic spongiform encephalopathy (BASE) is a newly recognized form of bovine prion disease, which was originally detected in Italy in 2004 as an effect of active surveillance. BASE or BSE L-type (L is referred to the lower electrophoretic PrPSc migration than classical BSE) has now been reported in several countries, including Japan. All field cases of BASE were older than 8 years and neurologically normal at the time of slaughtered. By experimental transmission, we defined the disease phenotype of cattle BASE, which is quite distinct from that seen in typical BSE and characterized by mental dullness and amyotrophy. Surprisingly, following intraspecies and interspecies transmission the incubation period of BASE was shorter than BSE. The relatively easy transmission of BASE isolate as well as the molecular similarity with sporadic Creutzfeldt-Jakob disease (sCJD) have raised concern regarding its potential passage to humans. Tg humanized mice Met/Met at codon 129 challenged with both BSE and BASE isolates, showed a resistance to BSE but a susceptibility to BASE at a 60% rate; in addition, BASE-inoculated Cynomolgus (129 Met/Met) had shorter incubation periods than BSE-inoculated primates. In this study we compared the biochemical properties of PrPSc in Cynomolgus and in TgHu Met/Met mice challenged with BSE and BASE strains, by conventional SDS-PAGE analysis and 2D separation. The results obtained disclose distinct conformational changes in PrPSc, which are dependent on the inoculated host but not on the codon 129 genotype.


 This work was supported by Neuroprion contract n. FOOD CT 2004 -506579 (NOE)



 WE know now, and we knew decades ago, that 5.5 grams of suspect feed in TEXAS was enough to kill 100 cows.


 look at the table and you'll see that as little as 1 mg (or 0.001 gm) caused 7% (1 of 14) of the cows to come down with BSE;


 Risk of oral infection with bovine spongiform encephalopathy agent in primates


 Corinne Ida Lasmézas, Emmanuel Comoy, Stephen Hawkins, Christian Herzog, Franck Mouthon, Timm Konold, Frédéric Auvré, Evelyne Correia, Nathalie Lescoutra-Etchegaray, Nicole Salès, Gerald Wells, Paul Brown, Jean-Philippe Deslys Summary The uncertain extent of human exposure to bovine spongiform encephalopathy (BSE)--which can lead to variant Creutzfeldt-Jakob disease (vCJD)--is compounded by incomplete knowledge about the efficiency of oral infection and the magnitude of any bovine-to-human biological barrier to transmission. We therefore investigated oral transmission of BSE to non-human primates. We gave two macaques a 5 g oral dose of brain homogenate from a BSE-infected cow. One macaque developed vCJD-like neurological disease 60 months after exposure, whereas the other remained free of disease at 76 months. On the basis of these findings and data from other studies, we made a preliminary estimate of the food exposure risk for man, which provides additional assurance that existing public health measures can prevent transmission of BSE to man.




 BSE bovine brain inoculum


 100 g 10 g 5 g 1 g 100 mg 10 mg 1 mg 0·1 mg 0·01 mg


 Primate (oral route)* 1/2 (50%)


 Cattle (oral route)* 10/10 (100%) 7/9 (78%) 7/10 (70%) 3/15 (20%) 1/15 (7%) 1/15 (7%)


 RIII mice (ic ip route)* 17/18 (94%) 15/17 (88%) 1/14 (7%)


 PrPres biochemical detection


 The comparison is made on the basis of calibration of the bovine inoculum used in our study with primates against a bovine brain inoculum with a similar PrPres concentration that was inoculated into mice and cattle.8 *Data are number of animals positive/number of animals surviving at the time of clinical onset of disease in the first positive animal (%). The accuracy of bioassays is generally judged to be about plus or minus 1 log. ic ip=intracerebral and intraperitoneal.


 Table 1: Comparison of transmission rates in primates and cattle infected orally with similar BSE brain inocula


 Published online January 27, 2005



 It is clear that the designing scientists must also have shared Mr Bradley’s surprise at the results because all the dose levels right down to 1 gram triggered infection.



 it is clear that the designing scientists must have also shared Mr Bradleyâs surprise at the results because all the dose levels right down to 1 gram triggered infection.



 Comment from Terry Singeltary This is a Comment on the Food and Drug Administration (FDA) Notice: Draft Guidance for Industry on Ensuring Safety of Animal Feed Maintained and Fed On-Farm; Availability


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 Guidance for Industry Ensuring Safety of Animal Feed Maintained and Fed On-Farm Draft Guidance FDA-2014-D-1180 Singeltary Comment


 Greetings FDA et al,


 I wish to comment on Guidance for Industry Ensuring Safety of Animal Feed Maintained and Fed On-Farm Draft Guidance FDA-2014-D-1180.


 Once again, I wish to kindly bring up the failed attempt of the FDA and the ruminant to ruminant mad cow feed ban of August 4, 1997. This feed ban is still failing today, as we speak. Even more worrisome, is the fact it is still legal to feed cervids to cervids in the USA, in fact, the FDA only _recommends_ that deer and elk considered to be of _high_ risk for CWD do not enter the animal food chain, but there is NO law, its only voluntary, a recipe for a TSE prion disaster, as we have seen with the ruminant to ruminant feed ban for cattle, where in 2007, one decade post August 1997 mad cow feed ban, where in 2007 10,000,000 POUNDS OF BANNED BLOOD LACED MEAT AND BONE MEAL WHEN OUT INTO COMMERCE, TO BE FED OUT. Since 2007, these BSE feed ban rules have been breached time and time again. tons and tons of mad cow feed went out in Alabama as well, where one of the mad cows were documented, just the year before in 2006, and in 2013 and 2014, breaches so bad (OAI) Official Action Indicated were issued. those are like the one issued where 10 million pounds of banned blood laced meat and bone meal were fed out.


 What is the use of having a Guidance for Industry Ensuring Safety of Animal Feed Maintained and Fed On-Farm Draft Guidance FDA-2014-D-1180, if it cannot be enforced, as we have seen with a mandatory ruminant to ruminant feed ban?


 I strenuously once again urge the FDA and its industry constituents, to make it MANDATORY that all ruminant feed be banned to all ruminants, and this should include all cervids as soon as possible for the following reasons...




 In the USA, under the Food and Drug Administrations BSE Feed Regulation (21 CFR 589.2000) most material (exceptions include milk, tallow, and gelatin) from deer and elk is prohibited for use in feed for ruminant animals. With regards to feed for non-ruminant animals, under FDA law, CWD positive deer may not be used for any animal feed or feed ingredients. For elk and deer considered at high risk for CWD, the FDA recommends that these animals do not enter the animal feed system.


 ***However, this recommendation is guidance and not a requirement by law.




 31 Jan 2015 at 20:14 GMT


 *** Ruminant feed ban for cervids in the United States? ***


 31 Jan 2015 at 20:14 GMT





19 May 2010 at 21:21 GMT


*** Singeltary reply ; Molecular, Biochemical and Genetic Characteristics of BSE in Canada Singeltary reply ;



 Tuesday, December 23, 2014







Sunday, December 15, 2013





 DOCKET-- 03D-0186 -- FDA Issues Draft Guidance on Use of Material From Deer and Elk in Animal Feed; Availability


Date: Fri, 16 May 2003 11:47:37 0500


EMC 1 Terry S. Singeltary Sr. Vol #: 1









Date: March 21, 2007 at 2:27 pm PST




Blood meal used to make cattle feed was recalled because it was cross- contaminated with prohibited bovine meat and bone meal that had been manufactured on common equipment and labeling did not bear cautionary BSE statement.




42,090 lbs.








Products manufactured from bulk feed containing blood meal that was cross contaminated with prohibited meat and bone meal and the labeling did not bear cautionary BSE statement.




9,997,976 lbs.




ID and NV





 Terry S. Singeltary Sr.


*** See attached file(s)


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Guidance for Industry Ensuring Safety of Animal Feed Maintained and Fed On-Farm Terry Singeltary Comment View Attachment:



 Sunday, April 5, 2015


 *** Guidance for Industry Ensuring Safety of Animal Feed Maintained and Fed On-Farm Draft Guidance FDA-2014-D-1180 ***



 ‘’AS i said before, OIE should hang up there jock strap now, since it appears they will buckle every time a country makes some political hay about trade protocol, commodities and futures. IF they are not going to be science based, they should do everyone a favor and dissolve there organization.’’


 Wednesday, March 11, 2015


 OIE and Centers for Disease Control and Prevention Reinforce Collaboration



 Saturday, April 18, 2015


 vCJD TEXAS CDC Emerging Infectious Diseases May 2015 Baylor College of Medicine Neuroscience 2014 case of human form of “mad cow disease” highlights need for continued surveillance



 Saturday, May 09, 2015


 *** Psychiatric Symptoms in Patients With Sporadic Creutzfeldt-Jakob Disease in Germany ***



 Sunday, May 3, 2015





 Saturday, December 13, 2014


 Terry S. Singeltary Sr. Publications TSE prion disease


 Diagnosis and Reporting of Creutzfeldt-Jakob Disease


 Singeltary, Sr et al. JAMA.2001; 285: 733-734. Vol. 285 No. 6, February 14, 2001 JAMA







lost my mom to the Heidenhain Variant of Creutzfeldt Jakob Disease hvCJD 12/14/97 confirmed. I just made a promise to mom, never forget (I could never ever forget what I saw), and never let them forget...




 Terry S. Singeltary Sr., Bacliff, Texas USA 77518