Wednesday, July 28, 2010

Atypical prion proteins and IBNC in cattle DEFRA project code SE1796 FOIA Final report

Atypical prion proteins and IBNC in cattle DEFRA project code SE1796 FOIA Final report

Executive Summary

Studies of Bovine Spongiform Encephalopathy (BSE), carried out in the UK, showed it to be a single strain of prion disease based on histopathological (Simmons et al., 1996) and transmission data (Bruce et al., 1992 ). First reported in the 1980s (Wells et al., 1987) there appears to have been little change in the characteristics of the disease throughout the epidemic and BSE maintains a distinct molecular profile even following cross species transmission. However, during surveillance programmes in Europe and in North America two other distinct isolates of bovine prion disease have come to light, H and L type, so-called to reflect their unique molecular profiles (Yamakawa et al., 2003; Biacabe et al., 2004).

Reports were also emerging of atypical forms of scrapie that were distinct from classical scrapie isolates and were less easily recognised by the then current diagnostic tests (Benestad et al., 2003; Buschman et al., 2004). This led to concerns that cattle could also harbour a prion disease that was not detected by the current diagnostic tests for BSE. Importantly, approximately 15-20% of the clinical cases submitted for investigation were indeed negative and this proportion of negative cattle did not appear to vary despite increasing awareness of BSE clinical signs by the farming and veterinary community. While there maybe other explanations for this discrepancy (McGill et al., 1993), another underlying undiagnosed prion disease of cattle distinct from classical BSE could not be ruled out.

The study reported here investigated a small number of these BSE negative clinical cases by using more sensitive and modified diagnostic tests for abnormal PrP.

The majority of the cases that we studied were negative by all the tests employed and based on this observation we conclude that there was not a simultaneous epidemic of another form of bovine prion disease. However, we observed a number of classical cases that were missed prior to the advent of sensitive and rapid diagnostic tests and this provides an estimate of the number of cattle that were mis-diagnosed before 2000. In addition, we observed a few rare cases where the diagnostic tests were not in agreement and these cases were investigated further. One of these unusual samples emerged as a case of idiopathic bovine neuronal chromatolysis (IBNC).

During the study we also reported the first H-type BSE case in the UK (Terry et al., 2007).

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Scientific Objectives as prescribed in the project:

All of the objectives have been met and are described in detail below. Three annexes accompany this report, one with the figures for the results below and two papers for submission to peer-reviewed journals.

Objective 1:

To determine the variation of PK sensitivity of bovine PrPc from uninfected cattle brains and compare with bovine PrPsc from classical cases of BSE in order to set thresholds for negative, weak and strong positive values in commercially available rapid diagnostic tests.

Objective 2:

Determine whether there are a greater proportion of bovine brain samples positive for the rapid diagnostic tests (hereby called reactors) in the clinically-suspect, negative subset of cattle than in healthy negative cattle. (True positives will be determined on the basis of evaluation by IHC but should be strongly positive in both the rapid diagnostic tests).

Objective 3:

Determine whether the phenotypic and molecular characteristics of PrP from cattle identified in 2 are distinct from normal PrPc and from bovine PrPsc normally associated with classical BSE.

Studies of Bovine Spongiform Encephalopathy (BSE), carried out in the UK, showed it to be a single strain of prion disease based on histopathological (Simmons et al., 1996) and transmission data (Bruce et al., 1992). First reported in the 1980s (Wells et al., 1987) there appears to have been little change in the these characteristics of the disease throughout the epidemic; BSE also appear to maintain a distinct molecular profile in cattle and even when experimentally (or naturally) transmitted to other species such as humans and cats. However, during surveillance programmes in Europe, Japan and in North America, two other distinct isolates of bovine prion disease have come to light, H and L type, so-called to reflect their unique molecular profiles (Yamakawa et al., 2003; Biacabe et al., 2004).

In the late 1990’s, a novel prion disease was discovered in sheep (Benestad et al., 2003; Buschman et al., 2004); this Nor98 or atypical scrapie is widespread in Europe but had previously been missed by histopathological or immunohistological examination. This led to concerns that cattle could also harbour a prion disease that, unlike H- and L-type BSE, was not detected by the current diagnostic tests for BSE. Importantly, approximately 15-20% of the clinical cases submitted for investigation were indeed negative and this proportion of negative cattle did not appear to vary despite increasing awareness of BSE clinical signs by the farming and veterinary community. While there maybe other explanations for this discrepancy (McGill et al., 1993), another underlying undiagnosed prion disease of cattle distinct from classical BSE could not be ruled out.

The study reported here investigated a small number of these BSE negative clinical cases by using more sensitive and modified diagnostic tests for abnormal PrP. The majority of the cases that we studied were negative by all the tests employed and based on this observation we conclude that there was not a simultaneous epidemic of another form of bovine prion disease. However, we observed a number of cases of BSE in this “BSE negative” sub-set that were missed prior to the advent of more sensitive and rapid diagnostic tests and this provides an estimate of the number of cattle that were mis-diagnosed before 2000. In addition, we observed a few rare cases where the diagnostic tests were not in agreement and these cases were investigated further. One of these unusual samples emerged as a case of idiopathic bovine neuronal chromatolysis (IBNC) (Jeffrey & Wilesmith, 1992; 1996; Jeffrey et al., 2009).

During the study we also reported the first H-type BSE case in the UK (Terry et al., 2007).

Materials and Methods

Tissue samples.

Test samples: Frozen brain stem from 501 bovine BSE suspects with neurological signs, a) that were negative at the level of the obex for vacuolation by standard histopathological techniques from years 1991-1999 and b) by IHC and diagnostic Bio-Rad PlateliaTM from 2000 onwards. These tissues have been stored at the VLA at –80oC since submission.

Negative controls: Frozen brain stem from 90 cattle investigated as part of the active surveillance programme. These samples were submitted in 2006 to LGC for rapid testing by Bio-Rad TeSeE diagnostic ELISA and were negative. These samples were stored at –80oC prior to testing and were stored for a maximum of 36 months and therefore considerably less time than all experimental samples under investigation.

Cattle with suppurative encephalitis: 10 additional cattle samples were retrieved from the VLA Archive that were negative for BSE but showed signs of suppurative encephalitis and inflammation (lymphocyte cuffing and gliosis). These signs were consistent with listeria infection.

Tests for disease-associated PrP

IDEXX BSE Herdchek BSE antigen test kit

All samples were assayed using the IDEXX Herdchek Bovine Spongiform Encephalopathy (BSE) Antigen Test Kit, EIA according to the manufacturer’s instructions and without modification. Briefly, brains were homogenised in the buffer provided by the manufacturer and diluted prior to adding to the seprion (polyanion) coated multiwell plate and incubated prior to washing. The samples were then treated with a conditioning buffer to expose the antigen epitopes. PrPsc was detected by PrP specific antibodies conjugated to horseradish peroxidase and visualised with TMB substrate. Samples were read using a microtitre plate reader (Victor-Perkin-Elmer). The method has no Proteinase K digestion step and has only a mild trypsin treatment that is not required for specificity but aids in the epitope exposure step. The normal curve of negative control samples is provided by the manufacturer and shows the diagnostic cut off value is set higher than most negative controls. The amount of brain added to a single well is approximately 20 mg.

Diagnostic Bio-Rad TeSeE EIA

Sample extraction and detection was carried out according to the manufacturer’s instructions for the Bio-Rad TeSeE BSE ELISA. Briefly brain samples were homogenised in buffer provided by the manufacturer and then treated for 10 mins with Proteinase K at 37oC . The PK concentration is not provided by the manufacturer so we refer to it as 4 ul/ml which is the quantity of stock PK to final solution directed by Bio-Rad. A comparison with sigma PK indicated that the concentration is approximately 40 ug/ml. The samples were then precipitated and concentrated by centrifugation. Pellets were reconstituted and diluted in the buffers provided by the manufacturer. The PrPsc was then detected by a sandwich ELISA provided by the manufacturer. Details of the antibodies are not provided. Samples are read using a microtitre plate as above. Cut off values for the ELISA are calculated using the mean of four negative control ODs. The manufacturers indicate that a value of 0.14 should be added to the negative control mean and samples equal to or greater than this value should be further analysed. The amount of brain added to a single well is approximately 65 mg.

BioRad TeSeE EIA with reduced PK digestion (0.3 Bio-Rad TeSeE ELISA)

The PrPsc associated with atypical scrapie is believed to be less PK resistant than classical scrapie (Everest et al., 2006). In order to investigate whether an atypical form of BSE in cattle exists biochemically similar to atypical scrapie a modified version of the Bio-Rad TeSeE protocol, using sub-diagnostic levels of Proteinase K (0.3ul/ml), was used. This quantity of PK was arrived at by titration of PK and digestion of PrPc from 47 cattle brains negative for TSEs.

The Bio-Rad TeSeE BSE diagnostic test was used as directed by the manufacturer with the addition of DNAse prior to the Proteinase K (0.3 ul/ml PK) treatment and Pefabloc was added alongside the kit PK stopping solution. The PK dilution for these assays was prepared from a Sigma stock solution and 0.3 units/ml was the equivalent activity as 0.3 ul/ml of Biorad PK.

Bio-Rad TeSeE Western Blot

Sample extraction was carried out according to the manufacturer’s instructions (Bio-Rad TeSeE Western Blot) with several modifications. In brief, brain tissue was ribolysed to give 20 % (w/v) homogenate and subsequently incubated with DNAse. The samples were then digested with 0.3, 1, 4 or 20 units/ml PK (Sigma; where units/ml is an in-house nomenclature and 0.3 units/ml is equivalent to 0.3 µl of the Bio-Rad test PK in terms of activity as compared using the TAME test -Pierce) and the reaction stopped with Pefabloc. Following precipitation and centrifugation at 15,000 g for 7 minutes, in accordance with the Bio-Rad TeSeE Western blot protocol, the pellets were re-suspended in Laemmli sample buffer.

For analysis, the supernatants were heated at 100oC for 5 minutes, loaded on a 12% Criterion XT Bis-tris SDS gel (Bio-Rad) and subjected to electrophoresis in XT-MOPS running buffer (Bio-Rad) at 200 V for 50 minutes. Proteins were transferred to a PVDF membrane (Bio-Rad) at 115 V, 60 min using Tris/CAPS transfer buffer (Bio-Rad).

Blots to be evaluated using the Sha31 (Bio-Rad) antibody were incubated for one hour with the blocking solution provided by the manufacturers; and antibodies SAF84 (aa 175-180), P4 (aa 89-104) and FH11 (aa 55-65) using a 5% milk powder in PBS supplemented with Tween 20 (PBST). The membranes were incubated for one hour with the primary antibody and then with goat anti-mouse IgG antibody conjugated to horseradish peroxidase (Bio-Rad) prior to visualization by chemiluminescence (ECL; Amersham).

Immunohistochemical analysis

Formalin-fixed, paraffin wax-embedded tissue blocks were sectioned at 4mm, collected onto frosted charged slides (GmbH) and melted on at 60°C overnight to improve adhesion. Sections were de-waxed in xylene and alcohol and washing in water. They were subsequently put into 98% formic acid (Merck) for 30 minutes, washed in running tap water for 15 minutes and then fully immersed into citrate buffer (200mM trisodium citrate dehydrate (Sigma), 30mM citric acid (Sigma), pH 6.1) prior to being autoclaved for 30 minutes at 121°C. Endogenous peroxidase activity was quenched using 3% hydrogen peroxide (Sigma) and the sections immersed in purified water and stored at 4°C overnight. After warming to room temperature, non-specific antibody binding sites were blocked using normal goat serum (Vector Laboratories) for 20 minutes. Rat monoclonal anti-PrP R145 (VLA) was diluted to 2mg/ml and applied for one hour at ambient (19°C-24°C) temperature. Biotinylated rabbit-anti-mouse IgG (Vector Laboratories) was diluted appropriately and applied for 30 minutes at ambient temperature. Elite ABC (Vector Laboratories) was prepared according to the manufacturers’ directions and applied for 20 minutes at ambient temperature. Sections were washed between each stage using 5mM tris buffered saline supplemented with Tween-20 (5mM tris, 0.85% NaCl, 0.05% tween-20 (all from Sigma), pH 7.6). Diaminobenzidine tablets (Sigma) were prepared in McIlvanes buffer (200mM disodium hydrogen orthophosphate, 100mM citric acid (both from Sigma), pH 6.4) and applied for 10 minutes at ambient temperature. Sections were counterstained in Mayer’s haematoxylin and “blued” in running tap water, before being dehydrated through three changes each of absolute alcohol and xylene for three minutes each and finally mounted in DPX (Sigma).

Definition of terms

Disease associated isoforms of PrP may be distinguished from normal PrP by its increased resistance to Proteinase digestion in immunoblotting or ELISA tests (PrPres), binding to polyanions or labelling with PrP specific antibodies in fixed and treated paraffin-embedded section (PrPd). Included within the operational definition of PrPd are all those detection systems that do not use Proteinase K digestion. The correlation between prion infectivity and PrPres or PrPd is inexact, and infectivity has been dissociated from PrPres or PrPd in several experiments, putatively this is because only a fraction of abnormal PrP isoforms are infectious. We will therefore use operational definitions for detected abnormal PrP forms and PrPsc for the hypothetical infectious sub-population of PrP isoforms detected by bioassay.

Results

Brains from cattle previously diagnosed as negative for BSE based on histopathological examination were investigated in this study for evidence of unusual prion diseases. The majority of the cattle investigated were submitted to the VLA as BSE suspect during the years 1997-2005 and were reported to have clinical signs similar to BSE. We applied a combination of modified and previously unused diagnostic tests to this subgroup of cattle including lower concentrations of PK for protein digestion, tests that do not use PK for PrPsc detection and standard Western blot (WB) procedures with Mabs reactive with different regions of the PrP glycoprotein. A flow chart detailing the sequence for the investigation of potential unusual prion diseases of cattle are shown in Figure 1.

1) Determination of the lowest PK concentration that digests PrPc from brains of cattle

The minimum concentration of PK required for the elimination of PrPc in the majority of non-exposed control cattle samples, resulting in a negative value in the Bio-Rad TeSeE ELISA, was determined. PK titrations were performed on BSE positive and negative control reference material (CRM) and subsequently on 47 individual confirmed negative brainstems. The brainstems had previously tested negative with the diagnostic Bio-Rad TeSeE ELISA by LGC and were obtained from active surveillance and therefore unlikely to have had clinical signs of disease. An amount of 0.3 µl/ml PK was selected for use in the adapted Bio-Rad TeSeE ELISA (0.3 Bio-Rad) (Table 1).

2) Determination of threshold values for the IDEXX HerdChek and 0.3 modified Bio-Rad rapid tests

The diagnostic tests have cut-off values that are set by the manufacturers. For the 0.3 Bio-Rad ELISA new cut-off values were determined to take account of the modifications. While no modifications were made to the IDEXX HerdChek assay cut-off values were calculated using the same test samples for consistency. 90 confirmed BSE negative brainstems were assayed and threshold values calculated as 3 standard deviations above the mean (Table 2). Threshold values of 0.166 Absorbance Units (AU) and 0.137 AU were set for the 0.3 Bio-Rad TeSeE and IDEXX Herdchek EIAs respectively. A single confirmed negative sample gave a value above the IDEXX threshold limit (0.240AU) on first assay. However, when repeated this sample was negative (0.016AU).

3) Results of assays applied to the test BSE cattle population

The assays described above and mapped in Figure 1 were then applied to the brains from 501 clinically suspect cattle. Following analysis the cattle were divided into five groups and these are described below. The results are summarised in Table 3.

Group1: Confirmed negative diagnosis of clinically suspect cattle

Brainstems from 501 cattle submitted to the VLA for BSE diagnosis between the years 1991 and 2005 that were subsequently diagnosed as negative by the tests used at time of slaughter, were assayed using the IDEXX and 0.3 Bio-Rad immunoassays for detection of abnormal PrP. 436 (87%) were negative by both tests. All of the samples submitted after 1999 were confirmed negative (see below) (Figure 8). By these criteria we were unable to detect abnormal PrP in the brainstems of these cattle and this subset of clinically suspect cattle is unlikely to harbour a prion disease. However, we were unable to test other areas of the brain from these cattle and PrPsc distribution patterns distinct from classical BSE cannot be ruled out. In addition to the 501 brainstems we also tested 191 cerebella by the same methods, all of which were negative by standard tests.

Group 2: Confirmed positive for BSE by all diagnostic tests

Sixty five samples remained that were positive in either the IDEXX or the 0.3 Bio-Rad assays or in both of these tests. Of these, 40 were positive by both tests (modified as above) and following retesting were positive using diagnostic concentrations of PK for the Bio-Rad TeSeE (figure 2). Immunohistochemical evaluation of abnormal PrP in the obex demonstrated normal distribution of PrPsc deposits similar to those observed for classical BSE (Figure 8).

To confirm that the PK resistant glycoproteins of abnormal PrP resembled the molecular profile of classical BSE, all 40 cases were immunoblotted using SHa31 MAb (figure 3). In all cases a signature 3 glycoprotein banding pattern was observed with relative mass and glycoprotein ratios indistinguishable from classical BSE. These animals ranged in age from 5 years to 12 years, with a mean age of 6 years, 10 months. All 40 animals were female and comprised 32 Friesians, 2 Holsteins, 2 Herefords, 1 Limousin/Friesian Cross, 2 Holstein/Friesian Cross and 1 Simmental.

The 40 confirmed positive samples were from cattle slaughtered between the years 1997 and 1999. We tested a total of 285 from this period and this represents 14.0% of the clinical suspects that were confirmed negative for BSE at this time. If this is representative of the entire clinical suspect unconfirmed cattle (total 2,426) during this period (1997-1999 inclusive) a total of 340 BSE positive cattle would have been missed. This under-diagnosis is likely to be a result of the diagnostic tests applied at the time. Up until the year 2000, all BSE cases were diagnosed by detection of vacuolation and gliosis in the obex. It is clear that this method is not 100% sensitive for prion diseases either because not all cases present with vacuolation or that vacuolation is a late onset phenomenon during clinical disease (Arnold et al., 2007). Our data showed that there were no additional cases of under-diagnosis after more sensitive diagnostic tests were introduced in 2000. During the years 1997-1999, a total of 12,171 clinical cases were submitted for BSE diagnosis of which 9,745 (80.1%) were confirmed positive with an estimated 2.8% of the total suspects submitted under-diagnosed by our calculations.

Assuming no other factors influenced the levels of correct diagnosis and that the numbers estimated for 1997 to 1999 were a true representation of the potential under-diagnosis of the entire epidemic up until 1999, then the total number of missed cases positive for BSE could have been in the region of 5,500.

A draft version of this manuscript has been prepared.

Group 3: Confirmed positive for BSE by all rapid diagnostic tests but negative by IHC

2 of the 501 negative subset brainstem tested were positive by standard biochemical, diagnostic tests (Table 4) but abnormal PrP deposits were not observed in the obex when evaluated by IHC (Figure 8). This is clearly an unusual finding and both cases were rigorously audited prior to further investigation to determine that the sample for biochemistry was identical to the paraffin-embedded sample. As far as can be determined no errors in sampling and dispatch occurred for these two samples. Further DNA profiling and matching frozen sample to histology processed sample would confirm this. There was insufficient sample to perform any further analysis on one case, but the other case was further investigated using the modified TeSeE Western blot protocol described above – at the diagnostic standard PK concentration of 4 µl/ml for PrPsc digestion. Western blotting of abnormal PrP from this sample confirmed the ELISA data with intense labelling of PK resistant PrP using the PrP-specific antibodies Sha31 and SAF84 (Figure 4a and 4b). The glycoprofile and molecular mass of the PrP bands were indistinguishable from classical BSE A band was labelled strongly with FH11 Mab (that recognises an N terminal PrP epitope) and is therefore likely to represent undigested PrP (Figure 4c). In addition, at 4 µl/ml PK, strong reactivity is also observed with the P4 mAb (Figure 4d). Molecular comparison of this case with classical BSE and with scrapie – using different levels of PK, different dilutions of positive sample and different PrP-specific antibodies, indicates that there is no discernible difference of the test sample with classical BSE. Both cases were extensively followed up by IHC using Mabs to different regions of the PrP molecule but were negative in all cases (data not shown).

Why the PrPsc could not be detected by IHC is unclear. Further analysis by transmission to rodent models of prion disease may shed further light on the characteristics of this sample. Indeed, murine models of prion disease have been reported where PrPsc cannot be detected in the brains but these studies confirmed the lack of PrPsc by all assays including Western Blot.

Group 4: IDEXX Herdchek positive, 0.3 Bio-Rad negative, IHC positive.

Two brainstem samples (98/00819; 98/02316) were positive by the IDEXX Herdchek EIA (Table 5) but Bio-Rad test negative even following PK digestion at sub-optimal concentrations. Both of these samples demonstrated abnormal PrP deposition in the obex by IHC evaluation (Figure 8). Western blot analysis of PK resistant PrP glycoprotein from sample 98/2316 indicated that low amounts of PrPres could be detected using Sha31 and SAF84 Mabs. From these blots and taking into account the low levels of PrPres detected the banding patterns appeared indistinguishable from classical BSE (Figure 5a and 5b). No further sample was available for 98/00819.

The sample contained very low levels of PrPres as shown by the WB data and this is likely to be the reason for lack of signal in the Bio-Rad ELISA. At these levels of abnormal PrP we are at the threshold of detection. The IDEXX HerdChek assay has consistently shown a higher analytical sensitivity for classical scrapie in our hands than the Bio-Rad assays. The values for the IDEXX HerdChek were in the region of 0.15-0.88 and these values are much lower than any of the other samples we have tested in this study. These data suggest that the IDEXX assay is more analytically sensitive than the Bio-Rad TeSeE for BSE.

However, there are alternative explanations for the discordance in test results. The Bio-Rad TeSeE ELISA detects PrPres with Mabs that detect 2 regions of the molecule. Any changes in PrP sequence in the region of Mab binding could alter analytical sensitivity. Therefore the bovine PrP open reading frame from 98/02316 was compared with that of two classical BSE samples, all three samples were 6:6 with respect to the octapeptide repeat. The only mutation seen in this unusual sample was at codon 78 and this is a “silent” mutation in that it does not affect the PrP protein sequence (glutamine, Q78). The Western blot results suggest that the PK cleavage sites of sample 98/02316 were not different from classical cases of BSE. Therefore we conclude that PrPres concentration in this sample was low, as indicated by the control BSE positive brain homogenate, when diluted to a level of 1/250, still producing bands of a far greater density than the test sample when assayed neat.

Group 5: Diagnostic Bio-Rad and IDEXX negative, IHC negative but 0.3 Bio-Rad positive

Twenty-one of the clinical suspect brainstems tested by 0.3 Bio-Rad modified protocol had OD values above the calculated cut off point (range 0.166 to 0.857) (Figure 6) but were IDEXX Herdchek negative and IHC negative (figure 8). The samples were also diagnostic Bio-Rad TeSeE negative and the cattle, all female, ranged in age from 3 years to 11.5 years. They comprised Friesian, Friesian/Holsten, Hereford Cross, Aberdeen Angus Cross, Simmental Cross and Limousin Cross breeds. These samples, where sufficient tissue was available, were analysed, for the presence of partially PK resistant PrP, using the Bio-Rad Western blot protocol with digestion carried out at 20 and 0.3 µl/ml of PK and detected using the SHa31 Mab. Following digestion of the samples with 20 µl/ml PK the samples were shown to be negative for the characteristic PrPsc banding patterns when compared to three individual BSE-negative samples and a classical BSE positive sample (Fig 7a). However, faint bands were observed at approximately 16 and 25 KDa for 4 of the samples (T5, T8-T10) but this faint banding is consistent with partially digested PrPc but could also be a result of variable amounts of protein loaded per lane. At 0.3 µl/ml PK, banding is observed for all test samples, with banding consistent with partially digested PrPC, as also observed for the three known BSE-negative samples. In contrast, the classical BSE-positive sample gave a distinct banding pattern, different from that observed for the BSE-negative samples (Fig.7b). Consistent with the above results samples T5 and T8-T10 demonstrated increased intensity of labelling that could result from an up-regulation or increase in PrPc and could also account for the high signals in the modified ELISA.

Variable banding intensity between lanes may also be a result of inconsistent loading of amounts of protein per lane. However, our previous experience of testing protein concentrations PRIOR to PK digestion in the individual samples showed that they were very consistent to within <5% of the total amount. In addition, although we add pefabloc to stop PK digestion it is also likely that there is variation in the PK digestion amongst samples. Both variables could account for the differences in intensities between lanes. However, we cannot exclude the possibilitity that a PK sensitive variant of abnormal PrP is present as demonstrated by Barron et al 2007 who also demonstrated a 22 KDa band following sub optimal PK digestion. The samples were further investigated as below. Encephalitis may up regulate PrPc One explanation for high values in the immunoassay following digestion with suboptimal concentrations of PK could be high levels of PrPc in the sample. Increased levels of PrPc may occur as a result of up-regulation of PrPc on tissue resident cells or from the influx of inflammatory cells into the site following infection. Differential diagnoses were available for 9 of the 21 animals and nine had confirmed encephalitic lesions and inflammation. Further to this observation we therefore analysed brainstems from 10 BSE negative cattle (but also clinical suspects) by both modified rapid tests that had confirmed encephalitis. The brainstems from 9 encephalitis cattle were negative by both the 0.3 Bio-Rad TeSeE and IDEXX assays. The brainstem from 1 animal was positive by the 0.3 Bio-Rad assay but negative by the IDEXX EIA. The result from this sample is similar to the 21 observed above in group 5. It is unclear therefore whether the high levels of PrP are a result of concurrent infection as there is not a 100% correlation. However, PrPc is more susceptible to endogenous proteases and a low signal could be partly explained by inappropriate handling of the tissue at post-mortem. Loss of PrP detection following retesting of group 5 samples. When all 21 samples were re-analysed from a fresh piece of tissue from the archive (likely to have been frozen and thawed by the archive staff) only one retested as positive (figure 6). Further analysis of this sample (sample number 99/00514) by Western blot has not shown any bands suggesting the presence of an atypical form of prion protein. Any PK sensitive PrP, whether PrPc or unusual prion disease-associated PrP, is likely to be affected by tissue handling techniques including freezing, thawing and the amount of time in storage. This could explain loss of signal. These samples may also represent a small number of outliers in the negative population. This is still higher than we would expect given that only 1/90 negative control samples were outliers in the original testing. Identification of Idiopathic Brainstem Neuronal Chromatolysis (IBNC) in group 5 samples One of the 21 samples identified in group 5 was shown to have IBNC following histological investigation (03/00002) (figure 8). Concurrently, we investigated the PrP distribution in known cases of IBNC (Jeffrey et al 2008; “Idiopathic Brainstem Neuronal Chromatolysis (IBNC): a novel prion protein related disorder of cattle?” BMC Vet Res. 2008 Sep 30;4:38. The IHC and histology profile of this case was very similar to that of the known IBNC cases. Investigation of the distribution and molecular characteristics of PrP from known IBNC See also: Idiopathic Brainstem Neuronal Chromatolysis (IBNC): a novel prion protein related disorder of cattle? Jeffrey M, Perez BB, Martin S, Terry L, González L. BMC Vet Res. 2008 Sep 30;4:38 Further investigations demonstrated that 57% the assays performed on the confirmed IBNC samples, using the 0.3 Bio-Rad TeSeE assay (n=42), gave values above those of the test kit control and also the BSE negative brain pool control. Half brains from six IBNC affected animals were retrieved from the TSE archive alongside the brainstem from a seventh animal. The cortex, brainstem, cerebellum and midbrain from these brains were sub-sampled and the adapted Bio-Rad TeSeE EIA, IDEXX Herdchek and Western Blot protocols applied to these tissues, in order to determine whether they could represent a form of atypical BSE. These samples had previously been found to be negative using the commercial Bio-Rad EIA and re-testing using this assay and the IDEXX Herdchek assay confirmed their negative status. When assayed using the adapted Bio-Rad protocol at 0.3µl/ml PK, 24/42 (57%) of the sample assays performed gave values above those of the test kit control and also the BSE negative brain pool control. Values above twice that of the calculated cut-off levels were found for each case but not for each brain site No PrPres was detected when Western blotting these samples at either 20 or 4µl/ml PK but a signal was detected on the gels when blotted at the 0.12 and 0.3µl/ml PK levels. At 0.12µl/ml PK the IBNC samples were indistinguishable from the negative controls but at the 0.3µl/ml level more PrPres was detected in the IBNC cases than in the controls with each of the antibodies tested (SHA31, F99, SAF84 and P4). Illustrations of the F99 blot are shown in the paper. Other data not shown. These data suggest that IBNC affected cattle abnormally express or accumulate PrP in brain and that the abnormal PrP is not strongly resistant to protease digestion. The results suggest that either the range of prion diseases is still wider than previously thought or that abnormalities of prion protein expression may be associated with brain lesions unconnected with prion disorders. Biochemical and transmission studies are planned in order to investigate further (under SE2014). First case of H-type BSE identified in GB During the course of this study, 1/5 frozen brainstem from bovine BSE cases when immunoblotted using the Bio-Rad TeSeE Western blot with antibodies P4, L42, 6H4, Sha31 and SAF84, was found to have a PrP profile indistinguishable from French H-type BSE. This sample was the first case of H-type BSE to be identified in GB. It was a fallen 13-year-old Galloway cow, first tested and confirmed as a case of BSE in November 2005. Due to autolysis its brain was unsuitable for further characterisation by IHC. Its age and reported absence of clinical signs are consistent with other cases of H-type BSE. When blotting the samples, mAbs Sha31 and 6H4 revealed, in this sample, an unglycosylated band with relative mobility less than BSE, and mAb P4, labelled the sample more strongly than the BSE samples hence supporting the observed similarities with the French H-type sample. Additionally, this study revealed in both this unusual sample and the French H-type a lower molecular weight band with relative mobility of between 6 and 10 kD labelled with the P4 and L42 mAbs. This band is not seen in BSE samples. This data was published in June 2007 (L. A. Terry et al. Veterinary record (2007) 160, 873-875). Discussion and Conclusions Here we report the investigation of 501 cattle samples that were submitted to the VLA for BSE diagnosis but subsequently confirmed as negative by the diagnostic test used at the time of submission. Prior to 2000 this was by histology alone and positive diagnosis was made solely upon the observation of vacuolation and gliosis in the relevant brain regions. As a result, using more sensitive diagnostic assays, we were able to diagnose BSE positive cattle from the years 1997-1999 inclusive that were originally negative by vacuolation. From these data we have estimated that approximately 3% of the total suspect cases submitted up until the year 1999 were mis-diagnosed. This is likely to be due to the relative sensitivities of the methods. In addition, it has been demonstrated in cattle that vacuolation occurs after PrPsc can be detected in the brain stem and that PrPsc is detected prior to clinical disease (Arnold et al, 2007). Thus these cattle may have suffering very early clinical signs. However, we have not ruled out the possibility that there may be a subset of BSE affected cattle where vacuolation at the obex does not occur. The two cattle that were positive by the rapid biochemical tests but negative by IHC is an unexplained observation. The samples both contained high amounts of abnormal prion protein as determined by the OD values from the rapid tests that according to our experience of confirmatory testing should have been easily detected by IHC. Furthermore, epitope mapping of the PK cleaved proteins demonstrated no unusual glycoform patterns and IHC evaluation with the same antibodies still did not reveal PrPd deposition in the wax embedded sections. Thus it is unlikely that lack of detection by IHC is the result of an unusual conformation of the PrPd that masks the epitope of R145, the antibody of choice for IHC evaluation at the VLA. The two cattle that were positive by all tests except Bio-Rad ELISA are easier to explain. Previously we have demonstrated that the IDEXX HerdChek scrapie antigen EIA is more analytically sensitive for scrapie than the Bio-Rad ELISA (project SE2007) and this also appears to be the case for bovine BSE. Indeed the two samples were positive by the Bio-Rad Western blot but with significantly reduced signals compared to a bovine positive control. Samples in group 5 were only positive in the Bio-Rad ELISA and only if sub-optimal concentrations of PK were used. Several explanations could account for this result. First, the samples may contain a subset of PrP molecules that have a slightly higher resistance to PK digestion than normal PrPc and that it is not sufficiently aggregated to be detected by the IDEXX assay; whether this is related to a prion disease or some other event that confers such properties on normal PrP remains unanswered. There are notable descriptions in the literature of TSE models where disease is not accompanied by the characteristic accumulation of PK resistant PrP or was found at extremely low levels (Piccardo et al., 2007; Barron et al 2007; Nazor et al., 2005). These findings together might suggest an additional family of neurodegenerative diseases where the infectious form of PrP is not readily detected by our current diagnostic tests. Second, the higher signal could be the result of an increase in the overall amount of PrPc in the samples as discussed in the results and related to up-regulation of PrP in cells resident in the brain or due to influx of inflammatory cells either as a result of damage or the presence of a non-prion related disease. Third, that the PrP in these samples is bound to an unidentified molecule that confers higher PK resistance, or fourth, inhibits proteinase K. IBNC is likely to represent a subset of this group of cattle. Based on these data, our overall conclusion is that a second type of BSE is unlikely to have co-existed at a high prevalence with the classical form in the cattle population during the UK epidemic.


snip...see full text ;


http://randd.defra.gov.uk/Document.aspx?Document=SE1796_8548_FIN.doc



IBNC BSE TSE update ;


Tuesday, November 17, 2009 SEAC NEW RESULTS ON IDIOPATHIC BRAINSTEM NEURONAL CHROMATOLYSIS (IBNC) FROM THE VETERINARY LABORATORIES AGENCY (VLA) SEAC 103/1


http://bse-atypical.blogspot.com/2009/11/seac-new-results-on-idiopathic.html


31 March 2009 - A summary of the 102nd SEAC meeting (35 KB) held on 4th March 2009


snip...


SEAC noted that IBNC appeared to be a rare disease that occurred in older cattle, predominantly as single cases, although it is possible that surveillance may not detect all cases. Biochemical studies suggested that the prion protein may play a role in the disease. However, it is unclear whether the normal form of the protein or an abnormal form is involved. Studies are required to determine whether IBNC is transmissible or not. SEAC concluded, noting that specified risk material controls are in place to prevent cattle brain from entering the food supply, that current data on IBNC do not suggest it presents a risk to human health.



http://www.seac.gov.uk/summaries/seac102_summary.pdf



>>> All of the 15 cattle tested showed that the brains had abnormally accumulated prion protein. <<<


Saturday, February 28, 2009

NEW RESULTS ON IDIOPATHIC BRAINSTEM NEURONAL CHROMATOLYSIS "All of the 15 cattle tested showed that the brains had abnormally accumulated PrP" 2009

SEAC 102/2

http://bse-atypical.blogspot.com/2009/02/new-results-on-idiopathic-brainstem.html



Wednesday, October 08, 2008

Idiopathic Brainstem Neuronal Chromatolysis (IBNC): a novel prion protein related disorder of cattle?

http://bse-atypical.blogspot.com/2008/10/idiopathic-brainstem-neuronal.html



8. I was in receipt of no extra funds beyond those provided by the NHS and the University of London to run my laboratories and pay my salary as a senior lecturer/honorary Consultant and I suffered no constraints over my publications, lectures to my students, or statements to the media. However, I became increasingly aware after 1988 that questioning official dogma about BSE brought difficulties to one’s career. I was myself about to retire from the Charing Cross Hospital, where I worked as a Consultant Neuropathologist, but I observed with horror that the good reputations of dissenting scientists in the field, not least Dr Stephen Dealler and especially Dr Harash Narang were systematically undermined.

http://collections.europarchive.org/tna/20080102135133/http://www.bseinquiry.gov.uk/files/ws/s410.pdf



THEY KNEW 2 DECADES AGO the damn BSE mad cow testing were not finding cases ;


BSE-NON-CONFIRMATION OF DISEASE

3. A question posed by Mr Whaley (para 2) is that classical lesions of BSE may not occur in all cases. Supposing we had a strain variant that produced it's lesions in the cerebrum these would not be detected by our current method. I think this would be unlikely but not impossible - another reason why at least a proportion of complete brains (or blocks) should be retained during the epidemic so if the problem Mr Whaley indicates escalates, it can be investigated.

snip...

5. IF you had the information what benefit would there be ? what would you do with it ?

CONCLUSION

I do not recommend any action. The situation should be accepted. I do not think the VIS can do more at present. The situation should be kept under review particularly if there is an escalation in numbers in this category.

R BRADLEY

15 MAY 1990

90/5.15/3.2

http://collections.europarchive.org/tna/20090505194948/http://bseinquiry.gov.uk/files/yb/1990/05/15003001.pdf



''THE LINE TO TAKE'' ON IBNC $$$ 1995 $$$

1995

page 9 of 14 ;

30. The Committee noted that the results were unusual. the questioned whether there could be coincidental BSE infection or contamination with scrapie. Dr. Tyrell noted that the feeling of the committee was that this did not represent a new agent but it was important to be prepared to say something publicly about these findings. A suggested line to take was that these were scientifically unpublishable results but in line with the policy of openness they would be made publicly available and further work done to test their validity. Since the BSE precautions were applied to IBNC cases, human health was protected. Further investigations should be carried out on isolations from brains of IBNC cases with removal of the brain and subsequent handling under strict conditions to avoid the risk of any contamination.

31. Mr. Bradley informed the Committee that the CVO had informed the CMO about the IBNC results and the transmission from retina and he, like the Committee was satisfied that the controls already in place or proposed were adequate. ...

snip... see full text ;


http://collections.europarchive.org/tna/20080102204938/http://www.bseinquiry.gov.uk/files/yb/1995/06/21005001.pdf




SEAC MINUTES OF THE 19TH MEETING HELD ON 21 JUNE 1995 AT THE CENTRAL VETERINARY LABORATORY

31. Mr Bradley informed the Committee that the CVO had informed the CMO about the IBNC results and the transmission from retina and he, like the Committee was satisfied that the controls already in place or proposed were adequate. ...

http://web.archive.org/web/20030327015011/http://www.bseinquiry.gov.uk/files/yb/1995/06/21005001.pdf




ALSO, please note, atypical BSE-H strain is also transmissible in the humanized transgenic mice with distinct phenotype ;


P26

TRANSMISSION OF ATYPICAL BOVINE SPONGIFORM ENCEPHALOPATHY (BSE) IN HUMANIZED MOUSE MODELS

Liuting Qing1, Fusong Chen1, Michael Payne1, Wenquan Zou1, Cristina Casalone2, Martin Groschup3, Miroslaw Polak4, Maria Caramelli2, Pierluigi Gambetti1, Juergen Richt5*, and Qingzhong Kong1 1Department of Pathology, Case Western Reserve University, Cleveland, OH 44106, USA; 2CEA, Istituto Zooprofilattico Sperimentale, Italy; 3Friedrich-Loeffler-Institut, Germany; 4National Veterinary Research Institute, Poland; 5Kansas State University, Diagnostic Medicine/Pathobiology Department, Manhattan, KS 66506, USA. *Previous address: USDA National Animal Disease Center, Ames, IA 50010, USA

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). Two atypical BSE strains, BSE-L (also named BASE) and BSE-H, have been discovered in three continents since 2004. The first case of naturally occurring BSE with mutated bovine PrP gene (termed BSE-M) was also found in 2006 in the USA. The transmissibility and phenotypes of these atypical BSE strains/isolates in humans were unknown. We have inoculated humanized transgenic mice with classical and atypical BSE strains (BSE-C, BSE-L, BSE-H) and the BSE-M isolate. We have found that the atypical BSE-L strain is much more virulent than the classical BSE-C. The atypical BSE-H strain is also transmissible in the humanized transgenic mice with distinct phenotype, but no transmission has been observed for the BSE-M isolate so far.

III International Symposium on THE NEW PRION BIOLOGY: BASIC SCIENCE, DIAGNOSIS AND THERAPY 2 - 4 APRIL 2009, VENEZIA (ITALY)

http://www.istitutoveneto.it/prion_09/Abstracts_09.pdf



BUT yet, USDA scientist even managed to change that science around too, by having the only cow known in the world to date that is familial BSE $ PLEASE NOTE, this so-called BSE with mutated bovine PrP gene (termed BSE-M) that was found in 2006 in the USA. a supposedly new strain of familial BSE ? takes me back to the infamous sporadic FFI, that's not familial ? they don't have a clue, in my opinion. but yet the USDA officials will blame it on anything and everything, but the most likely cause i.e. MAD COW FEED IN COMMERCE. Instead, now we have another new strain of mad cow disease, only this is in the USA, and it is just a spontaneous old cow disease i.e. 'familial h-BSEalabama'.


''We hypothesize that the bovine Prnp E211K mutation most likely has caused BSE in “the approximately 10-year-old cow” carrying the E221K mutation.''


WHAT a hoot. They have now taken the same strain of mad cow disease (h-BSE), that Kong et al in 2009 showed was transmissible to humans via human TG mice, and termed it another new mad cow disease, termed ‘‘U.S. BSE Alabama’’ as being another spontaneous happening from nothing. I swear, this just get's better and better. what about IBNC BSE, no cases yet in the USA ? and just what is IBNC BSE ? (more on that later).


BSE Case Associated with Prion Protein Gene Mutation

Bovine spongiform encephalopathy (BSE) is a transmissible spongiform encephalopathy (TSE) of cattle and was first detected in 1986 in the United Kingdom. It is the most likely cause of variant Creutzfeldt-Jakob disease (CJD) in humans. The origin of BSE remains an enigma. Here we report an H-type BSE case associated with the novel mutation E211K within the prion protein gene (Prnp). Sequence analysis revealed that the animal with H-type BSE was heterozygous at Prnp nucleotides 631 through 633. An identical pathogenic mutation at the homologous codon position (E200K) in the human Prnp has been described as the most common cause of genetic CJD. This finding represents the first report of a confirmed case of BSE with a potential pathogenic mutation within the bovine Prnp gene. A recent epidemiological study revealed that the K211 allele was not detected in 6062 cattle from commercial beef processing plants and 42 cattle breeds, indicating an extremely low prevalence of the E211K variant (less than 1 in 2000) in cattle.

Author Summary Top

Bovine spongiform encephalopathy (BSE or Mad Cow Disease), a transmissible spongiform encephalopathy (TSE) or prion disease of cattle, was first discovered in the United Kingdom in 1986. BSE is most likely the cause of a human prion disease known as variant Creutzfeldt Jakob Disease (vCJD). 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.


http://www.plospathogens.org/article/info%3Adoi%2F10.1371%2Fjournal.ppat.1000156



http://www.plospathogens.org/article/fetchObjectAttachment.action?uri=info%3Adoi%2F10.1371%2Fjournal.ppat.1000156&representation=PDF



WHAT does this study show. IS it really a novel mutation in the bovine prion protein gene (Prnp), called E211K, of a confirmed BSE positive cow from Alabama, United States of America, the only one in the world ?

How can scientist around the globe continue to believe such junk science coming from the ARS and USDA, and not see through all this, and still continue to want to trade with them ?

maybe I am not suppose to understand all this, or maybe they just wish I couldn't (and I am not claiming to understand all of it). I just know that something is not computing anymore with the UKBSEnvCJD theory, the rest of different atypical TSE, and the USDA and OIE only theory we are suppose to believe in. I see now where Japan and the USA are going to swap out BSE risk for FMD risk, and it's all about trade once again $ I have seen the FDA claim that feeding 5.5 grams of banned mad cow feed at the Purina plant in Gonzales Texas to each cow was not enough of the TSE agent to worry about. I have seen the TDAH and or the USDA et al completely cover up a highly suspect stumbling and staggering mad cow, sent straight to be rendered without any test at all for mad cow disease and then just claim ignorance, but we knew it was ordered from higher up officials. then we watched another mad cow in Texas, that was confirmed by a secret test to be a positive test, but yet this cow was ordered to be negative, until it took an act of Congress, and 7 months later and scientist complaining from around the globe, that this cow should be re-tested. IT was and it was CONFIRMED 7 months later. I have seen, and it was proven that part of the infamous June 2004 enhanced BSE surveillance and testing program was a sham, they were getting healthy brains from cattle they knew did not have mad cow disease and submitting them for testing. The were busted for that too. It was proven to be true. Then another 9,200 cows in the same program used a test least likely to find mad cow disease i.e. the IHC. oh yea, they did everything they could to claim the Washington cow that was positive, was not a USA cow, and in the end it worked, the suspect mad cow that was suppose to be from the USA, well that was not the right cow, it was another one, this one from Canada. after the back to back h-familial-BSE in Alabama, and h-BSE(whatever they come up with later) in Texas, right after these two cows were documented, they saw the writing on the wall and shut the testing down to numbers so low, it's now mathematically impossible to detect a mad cow case in the USA. what I have seen in these 13+ years is politics manipulate science, and it's not pretty.


The OIE and USDA et al sold there soul to the devil, and in doing so, they sold yours too.


EU IBNC BSE, ANOTHER OLD COW DISEASE, is m-BSE i.e. h-familial-BSE in Alabama only, is this EU IBNC BSE ???

OR, is this just another case of mad cow disease officials are trying to pawn off as something else $$$

Let's look at this closer.

h-familial-BSE in Alabama case

Obvious lesions of spongiform encephalopathy diagnostic for BSE were not present in the brainstem, however it was positive for the presence of PrPd by IHC (Figure 1B). Distribution of PrPd in the brainstem of this animal was not as uniform or as intense as seen with the C-type U.S. BSE case from 2003 (Figure 1C) [4]

Immunohistochemistry

Brain tissue was placed in 10% buffered formalin and after a minimum of 4 days of fixation appropriate sections of brainstem in the obex region were put in cassettes and kept in fresh formalin until they were processed for routine paraffin embedding. The procedure was described in detail previously [4]. The IHC results were interpreted as follows: (i) positive for PrPd: pink to red and (ii) background and negative for PrPd: only blue background. As positive controls, slides from the brainstem of a BSE-positive cow, obtained from the United Kingdom and from the U.S. BSE Case 2003 were used. As negative controls, slides from brainstem material of BSE-negative cattle and scrapie-negative sheep were used.

http://www.plospathogens.org/article/fetchObjectAttachment.action?uri=info%3Adoi%2F10.1371%2Fjournal.ppat.1000156&representation=PDF



IBNC BSE, that's not BSE, but is another novel prion disease.

Atypical prion proteins in cattle DEFRA project code SE1796 FOIA Final report

snip...

Investigation of the distribution and molecular characteristics of PrP from known IBNC

See also: Idiopathic Brainstem Neuronal Chromatolysis (IBNC): a novel prion protein related disorder of cattle?

Jeffrey M, Perez BB, Martin S, Terry L, González L.

BMC Vet Res. 2008 Sep 30;4:38

Further investigations demonstrated that 57% the assays performed on the confirmed IBNC samples, using the 0.3 Bio-Rad TeSeE assay (n=42), gave values above those of the test kit control and also the BSE negative brain pool control.

Half brains from six IBNC affected animals were retrieved from the TSE archive alongside the brainstem from a seventh animal. The cortex, brainstem, cerebellum and midbrain from these brains were sub-sampled and the adapted Bio-Rad TeSeE EIA, IDEXX Herdchek and Western Blot protocols applied to these tissues, in order to determine whether they could represent a form of atypical BSE. These samples had previously been found to be negative using the commercial Bio-Rad EIA and re-testing using this assay and the IDEXX Herdchek assay confirmed their negative status. When assayed using the adapted Bio-Rad protocol at 0.3µl/ml PK, 24/42 (57%) of the sample assays performed gave values above those of the test kit control and also the BSE negative brain pool control. Values above twice that of the calculated cut-off levels were found for each case but not for each brain site

No PrPres was detected when Western blotting these samples at either 20 or 4µl/ml PK but a signal was detected on the gels when blotted at the 0.12 and 0.3µl/ml PK levels. At 0.12µl/ml PK the IBNC samples were indistinguishable from the negative controls but at the 0.3µl/ml level more PrPres was detected in the IBNC cases than in the controls with each of the antibodies tested (SHA31, F99, SAF84 and P4). Illustrations of the F99 blot are shown in the paper. Other data not shown.

These data suggest that IBNC affected cattle abnormally express or accumulate PrP in brain and that the abnormal PrP is not strongly resistant to protease digestion. The results suggest that either the range of prion diseases is still wider than previously thought or that abnormalities of prion protein expression may be associated with brain lesions unconnected with prion disorders. Biochemical and transmission studies are planned in order to investigate further (under SE2014).

http://randd.defra.gov.uk/Document.aspx?Document=SE1796_8548_FIN.doc



c-BSE, atypical l-BSE, atypical h-BSE, m-BSE and or atypical familial h-BSEalabama, IBNC

just what is this IBNC and or the m-BSE in the bovine, and or could they be the same ?

are they just another strain of BSE ?


TSS

Labels: , , , , ,

Sunday, June 07, 2009

L-TYPE-BSE, H-TYPE-BSE, C-TYPE-BSE, IBNC-TYPE-BSE, TME, CWD, SCRAPIE, CJD, NORTH AMERICA

L-TYPE-BSE, H-TYPE-BSE, C-TYPE-BSE, IBNC-TYPE-BSE, TME, CWD, SCRAPIE, CJD, NORTH AMERICA, something to ponder ;

confusious ask, what if ;


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)



http://www.istitutoveneto.it/prion_09/Abstracts_09.pdf




P26

TRANSMISSION OF ATYPICAL BOVINE SPONGIFORM ENCEPHALOPATHY (BSE) IN HUMANIZED MOUSE MODELS

Liuting Qing1, Fusong Chen1, Michael Payne1, Wenquan Zou1, Cristina Casalone2, Martin Groschup3, Miroslaw Polak4, Maria Caramelli2, Pierluigi Gambetti1, Juergen Richt5*, and Qingzhong Kong1

1Department of Pathology, Case Western Reserve University, Cleveland, OH 44106, USA; 2CEA, Istituto Zooprofilattico Sperimentale, Italy; 3Friedrich-Loeffler-Institut, Germany; 4National Veterinary Research Institute, Poland; 5Kansas State University, Diagnostic Medicine/Pathobiology Department, Manhattan, KS 66506, USA. *Previous address: USDA National Animal Disease Center, Ames, IA 50010, USA

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). Two atypical BSE strains, BSE-L (also named BASE) and BSE-H, have been discovered in three continents since 2004. The first case of naturally occurring BSE with mutated bovine PrP gene (termed BSE-M) was also found in 2006 in the USA. The transmissibility and phenotypes of these atypical BSE strains/isolates in humans were unknown.

We have inoculated humanized transgenic mice with classical and atypical BSE strains (BSE-C, BSE-L, BSE-H) and the BSE-M isolate. We have found that the atypical BSE-L strain is much more virulent than the classical BSE-C. The atypical BSE-H strain is also transmissible in the humanized transgenic mice with distinct phenotype, but no transmission has been observed for the BSE-M isolate so far.



http://www.istitutoveneto.it/prion_09/Abstracts_09.pdf



Wednesday, February 11, 2009

Atypical BSE North America Update February 2009 Both of the BSE cases ascertained in the US native-born cattle were atypical cases (H-type), which contributed to the initial ambiguity of the diagnosis. 174, 185 In Canada, there have been 2 atypical BSE cases in addition to the 14 cases of the classic UK strain of BSE2: one was the H-type, and the other was of the L-type.198


snip...end


source : Enhanced Abstract Journal of the American Veterinary Medical Association January 1, 2009, Vol. 234, No. 1, Pages 59-72 Bovine spongiform encephalopathy Jane L. Harman, DVM, PhD; Christopher J. Silva, PhD



http://avmajournals.avma.org/doi/ref/10.2460/javma.234.1.59




Atypical BSE North America Update February 2009



http://bse-atypical.blogspot.com/2009/02/atypical-bse-north-america-update.html



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: mhtml:%7B33B38F65-8D2E-434D-8F9B-8BDCD77D3066%7Dmid://00000110/!x-usc:mailto:emmanuel.comoy@cea.fr



http://www.plosone.org/article/info:doi/10.1371/journal.pone.0003017




Saturday, December 01, 2007

Phenotypic Similarity of Transmissible Mink Encephalopathy in Cattle and L-type Bovine Spongiform Encephalopathy in a Mouse Model

Volume 13, Number 12–December 2007 Research

Phenotypic Similarity of Transmissible Mink Encephalopathy in Cattle and L-type Bovine Spongiform Encephalopathy in a Mouse Model

Thierry Baron,* Anna Bencsik,* Anne-Gaëlle Biacabe,* Eric Morignat,* andRichard A. Bessen†*Agence Française de Sécurité Sanitaire des Aliments–Lyon, Lyon, France; and†Montana State University, Bozeman, Montana, USA

Abstract

Transmissible mink encepholapathy (TME) is a foodborne transmissible spongiform encephalopathy (TSE) of ranch-raised mink; infection with a ruminant TSE has been proposed as the cause, but the precise origin of TME is unknown. To compare the phenotypes of each TSE, bovine-passaged TME isolate and 3 distinct natural bovine spongiform encephalopathy (BSE) agents (typical BSE, H-type BSE, and L-type BSE) were inoculated into an ovine transgenic mouse line (TgOvPrP4). Transgenic mice were susceptible to infection with bovine-passaged TME, typical BSE, and L-type BSE but not to H-type BSE. Based on survival periods, brain lesions profiles, disease-associated prion protein brain distribution, and biochemical properties of protease-resistant prion protein, typical BSE had a distint phenotype in ovine transgenic mice compared to L-type BSE and bovine TME.The similar phenotypic properties of L-type BSE and bovine TME in TgOvPrP4 mice suggest that L-type BSE is a much more likely candidate for the origin of TME than is typical BSE.

snip...

Conclusion

These studies provide experimental evidence that the Stetsonville TME agent is distinct from typical BSE but has phenotypic similarities to L-type BSE in TgOvPrP4 mice. Our conclusion is that L-type BSE is a more likely candidate for a bovine source of TME infection than typical BSE. In the scenario that a ruminant TSE is the source for TME infection in mink, this would be a second example of transmission of a TSE from ruminants to non-ruminants under natural conditions or farming practices in addition to transmission of typical BSE to humans, domestic cats, and exotic zoo animals(37). The potential importance of this finding is relevant to L-type BSE, which based on experimental transmission into humanized PrP transgenic mice and macaques, suggests that L-type BSE is more pathogenic for humans than typical BSE (24,38).



http://www.cdc.gov/eid/content/13/12/1887.htm?s_cid=eid1887_e




Transmissible Mink Encephalopathy TME

Subject: In Confidence - Perceptions of unconventional slow virus diseasesof animals in the USA - APRIL-MAY 1989 - G A H Wells




http://transmissible-mink-encephalopathy.blogspot.com/2007/12/phenotypic-similarity-of-transmissible.html




Dr. Thornsberry: Let's hypothesize that I had some cattle on the eastern slope and they were in the same pasture with elk with CWD. If a cow had been exposed to the PRP Scrapie and it did develop disease four years later, would that look like BSE? Would there be a way to determine if it came from CWD?

Dr. Bartz: The IC studies in cattle indicate it does not look like BSE. The clinical signs of the IC/CWD cattle are more like downer cattle, and not aggressive. As far as finding the source of a bovine TSE, the gold standard is the lesion profile study where you take cattle tissue and inoculate it into mice with appropriate controls, wait until the mice come down, and do the lesion profiling.




http://transmissible-mink-encephalopathy.blogspot.com/



http://transmissible-mink-encephalopathy.blogspot.com/2006_12_01_archive.html




3.57 The experiment which might have determined whether BSE and scrapie were caused by the same agent (ie, the feeding of natural scrapie to cattle) was never undertaken in the UK. It was, however, performed in the USA in 1979, when it was shown that cattle inoculated with the scrapie agent endemic in the flock of Suffolk sheep at the United States Department of Agriculture in Mission, Texas, developed a TSE quite unlike BSE.339 The findings of the initial transmission, though not of the clinical or neurohistological examination, were communicated in October 1988 to Dr Watson, Director of the CVL, following a visit by Dr Wrathall, one of the project leaders in the Pathology Department of the CVL, to the United States Department of Agriculture.340 The results were not published at this point, since the attempted transmission to mice from the experimental cow brain had been inconclusive. The results of the clinical and histological differences between scrapie-affected sheep and cattle were published in 1995. Similar studies in which cattle were inoculated intracerebrally with scrapie inocula derived from a number of scrapie-affected sheep of different breeds and from different States, were carried out at the US National Animal Disease Centre.341 The results, published in 1994, showed that this source of scrapie agent, though pathogenic for cattle,

*** did not produce the same clinical signs of brain lesions characteristic of BSE. ***

3.58 There are several possible reasons why the experiment was not performed in the UK. It had been recommended by Sir Richard Southwood (Chairman of the Working Party on Bovine Spongiform Encephalopathy) in his letter to the Permanent Secretary of MAFF, Mr (now Sir) Derek Andrews, on 21 June 1988,342 though it was not specifically recommended in the Working Party Report or indeed in the Tyrrell Committee Report (details of the Southwood Working Party and the Tyrell Committee can be found in vol. 4: The Southwood Working Party, 1988–89 and vol. 11: Scientists after Southwood respectively). The direct inoculation of scrapie into calves was given low priority, because of its high cost and because it was known that it had already taken place in the USA.343 It was also felt that the results of such an experiment would be hard to interpret. While a negative result 337 Fraser, H., Bruce, M., Chree, A., McConnell, I. and Wells, G. (1992) Transmission of Bovine Spongiform Encephalopathy and Scrapie to Mice, Journal of General Virology, 73, 1891–7; Bruce, M., Chree, A., McConnell, I., Foster, J., Pearson, G. and Fraser, H. (1994) Transmission of Bovine Spongiform Encephalopathy and Scrapie to Mice: Strain Variation and the Species Barrier, Philosophical Transactions of the Royal Society of London, Series B, Biological Sciences, 343, 405–11 338 Bruce, M., Will, R., Ironside, J., McConell, I., Drummond, D., Suttie, A., McCordie, L., Chree, A., Hope, J., Birkett, C., Cousens, S., Fraser, H. and Bostock, C. (1997) Transmissions to Mice Indicate that ‘New Variant’ CJD is Caused by the BSE Agent, Nature, 389, 498–501 339 Clark, W., Hourrigan, J. and Hadlow, W. (1995) Encephalopathy in Cattle Experimentally Infected with the Scrapie Agent, American Journal of Veterinary Research, 56, 606–12 340 YB88/10.00/1.1 341 Cutlip, R., Miller, J., Race, R., Jenny, A., Katz, J., Lehmkuhl, H., Debey, B. and Robinson, M. (1994) Intracerebral Transmission of Scrapie to Cattle, Journal of Infectious Diseases, 169, 814–20 342 YB88/6.21/1.2 343 YB88/11.17/2.4 SCIENCE 84 would be informative, a positive result would need to demonstrate that when scrapie was transmitted to cattle, the disease which developed in cattle was the same as BSE.344 Given the large number of strains of scrapie and the possibility that BSE was one of them, it would be necessary to transmit every scrapie strain to cattle separately, to test the hypothesis properly. Such an experiment would be expensive. Secondly, as measures to control the epidemic took hold, the need for the experiment from the policy viewpoint was not considered so urgent. It was felt that the results would be mainly of academic interest.345 3.59 Nevertheless, from the first demonstration of transmissibility of BSE in 1988, the possibility of differences in the transmission properties of BSE and scrapie was clear. Scrapie was transmissible to hamsters, but by 1988 attempts to transmit BSE to hamsters had failed. Subsequent findings increased that possibility.



http://www.bseinquiry.gov.uk/pdf/volume2/chapter3.pdf




Monday, May 11, 2009

Rare BSE mutation raises concerns over risks to public health



http://bse-atypical.blogspot.com/2009/05/rare-bse-mutation-raises-concerns-over.html




Sunday, April 12, 2009 BSE MAD COW TESTING USA 2009 FIGURES Month Number of Tests

Feb 2009 -- 1,891

Jan 2009 -- 4,620



http://www.aphis.usda.gov/newsroom/hot_issues/bse/surveillance/ongoing_surv_results.shtml



SEE FULL TEXT ;



http://madcowtesting.blogspot.com/2009/04/bse-mad-cow-testing-usa-2009-figures.html




Monday, May 4, 2009

Back to the Past With New TSE Testing Agricultural Research/May-June 2009



http://madcowtesting.blogspot.com/2009/05/back-to-past-with-new-tse-testing.html




Sunday, May 10, 2009

Identification and characterization of bovine spongiform encephalopathy cases diagnosed and NOT diagnosed in the United States



http://bse-atypical.blogspot.com/2009/05/identification-and-characterization-of.html



Saturday, February 28, 2009NEW RESULTS ON IDIOPATHIC BRAINSTEM NEURONAL CHROMATOLYSIS TYPE BSE

"All of the 15 cattle tested showed that the brains had abnormally accumulated PrP" 2009
SEAC 102/2


http://bse-atypical.blogspot.com/2009/02/new-results-on-idiopathic-brainstem.html



Wednesday, October 08, 2008

Idiopathic Brainstem Neuronal Chromatolysis (IBNC): a novel prion protein related disorder of cattle?



http://bse-atypical.blogspot.com/2008/10/idiopathic-brainstem-neuronal.html




''THE LINE TO TAKE'' ON IBNC $$$ 1995 $$$

1995

page 9 of 14 ;30. The Committee noted that the results were unusual. the questioned whether there could be coincidental BSE infection or contamination with scrapie. Dr. Tyrell noted that the feeling of the committee was that this did not represent a new agent but it was important to be prepared to say something publicly about these findings. A suggested line to take was that these were scientifically unpublishable results but in line with the policy of openness they would be made publicly available and further work done to test their validity. Since the BSE precautions were applied to IBNC cases, human health was protected. Further investigations should be carried out on isolations from brains of IBNC cases with removal of the brain and subsequent handling under strict conditions to avoid the risk of any contamination.31. Mr. Bradley informed the Committee that the CVO had informed the CMO about the IBNC results and the transmission from retina and he, like the Committee was satisfied that the controls already in place or proposed were adequate. ...

snip... see full text


http://www.bseinquiry.gov.uk/files/yb/1995/06/21005001.pdf



Wednesday, August 20, 2008

Bovine Spongiform Encephalopathy Mad Cow Disease typical and atypical strains, was there a cover-up ?

August 20, 2008


snip...


another question, just how long have these atypical BSE TSEs been around in the bovine ???let's look at another case of atypical BSE in Germany way back in 1992 ;Subject: atypical BSE reported in 1992 and conviently slaughterd and incinerated and then swept under rug for about 12 years Date: April 26, 2007 at 1:08 pm PST 1992NEW BRAIN DISORDER3. WHAT ABOUT REPORTS OF NEW FORM OF BSE?THE VETERINARY RECORD HAS PUBLISHED AN ARTICLE ON A NEW BRAIN DISORDER OF CATTLE DISCOVERED THROUGH OUR CONTROL MEASURES FOR BSE. ALTHOUGH IT PRESENTS SIMILAR CLINICAL SIGNS TO BSE THERE ARE MAJOR DIFFERENCES IN THE HISTOPATHOLOGY AND INCUBATION PERIODS BETWEEN THE TWO. MUST EMPHASISE THAT THIS IS NOT BSE.4. IS THIS NEW BRAIN DISORDER A THREAT?WE DO NOT EVEN KNOW WHETHER THE AGENT OF THIS DISEASE IS TRANSMISSIBLE. IN ANY CASE, CASES SO FAR IDENTIFIED HAD SHOWN SIMILAR SYMPTOMS TO THOSE OF BSE, AND THEREFORE HAVE BEEN SLAUGHTERED AND INCINERATED, SO THAT IF A TRANSMISSIBLE AGENT WERE INVOLVED IT WOULD HAVE BEEN ELIMINATED. .......



http://www.bseinquiry.gov.uk/files/yb/1992/10/26001001.pdf




2. The Collinge/Will dispute appears to rumble on. Dr. Collinge had told Dr. Tyrrell that Dr. Will's response to his criticism about sharing material had been ''quite unacceptable'' (in spite of it's apparently conciliatory tone). Apparently Professor Allen was now going to try and arrange a meeting to resolve the dispute. No action here for MAFF, although Mr. Murray may be interested.3. Dr. Tyrrell regretted that the Committee had not seen the article on BBD. However he felt that for the time being NO specific action was called for. The most important need was to consider the possibility that the condition might be transmissible. As we have discussed, I suggested that we might circulate a paper to the members of the committee giving our appreciation of this condition (and perhaps of other non-BSE neurological conditions that had been identified in negative cases) and of any necessary follow up action. IF any Committee member felt strongly about this, or if the issue CAME TO A HEAD, we would call an interim meeting. He was happy with this approach. I would be grateful if Mr. Maslin could, in discussion with CVL and veterinary colleagues draft such a note, which will presumably very largely follow what Mr. Bradley's briefing paper has already said, taking account of DOH comments, We can then clear a final version with DOH before circulating it to Committee members.




http://www.bseinquiry.gov.uk/files/yb/1992/10/29005001.pdf




IN CONFIDENCE

This is a highly competitive field and it really will be a pity if we allow many of the key findings to be published by overseas groups while we are unable to pursue our research findings because of this disagreement, which I hope we can make every effort to solve.




http://www.bseinquiry.gov.uk/files/yb/1992/10/26002001.pdf




COLLINGE THREATENS TO GO TO MEDIA




http://www.bseinquiry.gov.uk/files/yb/1992/12/16005001.pdf




2. The discovery might indicate the existence of a different strain of BSE from that present in the general epidemic or an unusual response by an individual host.3. If further atypical lesion distribution cases are revealed in this herd then implications of misdiagnosis of 'negative' cases in other herds may not be insignificant.snip...This minute is re-issued with a wider distribution. The information contained herein should NOT be disseminated further except on the basis of ''NEED TO KNOW''.

R Bradley


http://www.bseinquiry.gov.uk/files/yb/1993/02/17001001.pdf



IN CONFIDENCE

BSE ATYPICAL LESION DISTRIBUTION


http://www.bseinquiry.gov.uk/files/yb/1993/03/14001001.pdf



ALABAMA MAD COW CASE



snip...


see full text ;



http://bse-atypical.blogspot.com/2008/08/bovine-spongiform-encephalopathy-mad.html



Friday, May 29, 2009

Characterization of a U.S. Sheep Scrapie Isolate with Short Incubation Time


http://scrapie-usa.blogspot.com/2009/05/characterization-of-us-sheep-scrapie.html


Friday, May 29, 2009

Seven Deer Test Positive for Chronic Wasting Disease During 2009 Spring Collections in Hampshire County, West Virginia



http://chronic-wasting-disease.blogspot.com/2009/05/seven-deer-test-positive-for-chronic.html



O.K. confusious asks, IF all these new atypical BSEs i.e. new strains of mad cow disease is just an 'OLD COW PRION DISEASE', why then can not the 'old human prion disease' such as the sporadic CJD, be from an 'old cow prion disease', same as the nvCJD 'young people mad cow disease' (which also happens in 74 year old), but why cannot the 'old cow prion diseases', i.e. l-BSE, h-BSE, and ibncBSE, cause the 'old people prion disease', which looks like sporadic CJD. seems that is what some of the pathology is showing ???

OH, that probably makes too much sense, and that the only answer could be that it's all just a happenstance of bad luck and or a spontaneous event, that just happens out of the clear blue sky $$$

IF this is the case, then where are all the SPONTANEOUS BSE CASES OF MAD COW DISEASE IN THE U.S.A., AND WHERE HAVE THEY BEEN BURIED IN THE USA OVER THE LAST 25 YEARS ???


Thursday, April 30, 2009

FDA Issues Final Guidance for Renderers on Substances Prohibited From Use in Animal Food or Feed CVM Update Back April 30, 2009



http://madcowfeed.blogspot.com/2009/04/fda-issues-final-guidance-for-renderers.html




Sunday, December 28, 2008

MAD COW DISEASE USA DECEMBER 28, 2008 an 8 year review of a failed and flawed policy



http://bse-atypical.blogspot.com/2008/12/mad-cow-disease-usa-december-28-2008-8.html




Wednesday, August 20, 2008

Bovine Spongiform Encephalopathy Mad Cow Disease typical and atypical strains, was there a cover-up ?



http://bse-atypical.blogspot.com/2008/08/bovine-spongiform-encephalopathy-mad.html





Monday, June 01, 2009

Biochemical typing of pathological prion protein in aging cattle with BSE



http://bse-atypical.blogspot.com/2009/06/biochemical-typing-of-pathological.html




SPORADIC CJD CASES RISING IN U.S.A


Monday, April 20, 2009 National Prion Disease Pathology Surveillance Center Cases Examined1 (December 31, 2008)

April 20, 2009

National Prion Disease Pathology Surveillance Center Cases Examined1 (December 31, 2008)

National Prion Disease Pathology Surveillance Center Cases Examined1

(December 31, 2008)

Year Total Referrals2 Prion Disease Sporadic Familial Iatrogenic vCJD

1996 & earlier 42 32 28 4 0 0

1997 115 68 59 9 0 0

1998 93 53 45 7 1 0

1999 115 69 61 8 0 0

2000 151 103 89 14 0 0

2001 210 118 108 9 0 0

2002 258 147 123 22 2 0

2003 273 176 135 41 0 0

2004 335 184 162 21 0 13

2005 346 193 154 38 1 0

2006 380 192 159 32 0 14

2007 370 212 185 26 0 0

2008 383 228 182 23 0 0

TOTAL 30715 17756 1490 254 4 2

1 Listed based on the year of death or, if not available, on year of referral; 2 Cases with suspected prion disease for which brain tissue and/or blood (in familial cases) were submitted; 3 Disease acquired in the United Kingdom; 4 Disease acquired in Saudi Arabia; 5 Includes 20 cases in which the diagnosis is pending, and 17 inconclusive cases; 6 Includes 25 cases with type determination pending in which the diagnosis of vCJD has been excluded.

Rev 2/13/09 National



http://www.cjdsurveillance.com/pdf/case-table.pdf



http://www.cjdsurveillance.com/resources-casereport.html



http://www.aan.com/news/?event=read&article_id=4397&page=72.45.45



*5 Includes 20 cases in which the diagnosis is pending, and 17 inconclusive cases; *6 Includes 25 cases with type determination pending in which the diagnosis of vCJD has been excluded.


Greetings,


it would be interesting to know what year these atypical cases occurred, as opposed to lumping them in with the totals only.

are they accumulating ?

did they occur in one year, two years, same state, same city ?

location would be very interesting ?

age group ?

sex ?

how was it determined that nvCJD was ruled out ?

from 1997, the year i started dealing with this nightmare, there were 28 cases (per this report), up until 2007 where the total was 185 cases (per this report), and to date 2008 is at 182. a staggering increase in my opinion, for something that just happens spontaneously as some would have us believe. i don't believe it, not in 85%+ of all sporadic CJD cases. actually, i do not believe yet that anyone has proven that any of the sporadic CJD cases have been proven to be a spontaneous misfolding of a protein. there are many potential routes and sources for the sporadic CJD's. ...TSS

please see full text here ;



http://prionunitusaupdate2008.blogspot.com/2009/04/national-prion-disease-pathology.html




Monday, June 01, 2009

Biochemical typing of pathological prion protein in aging cattle with BSE


http://bse-atypical.blogspot.com/2009/06/biochemical-typing-of-pathological.html



Friday, May 29, 2009

Characterization of a U.S. Sheep Scrapie Isolate with Short Incubation Time


http://scrapie-usa.blogspot.com/2009/05/characterization-of-us-sheep-scrapie.html




Friday, May 29, 2009

Seven Deer Test Positive for Chronic Wasting Disease During 2009 Spring Collections in Hampshire County, West Virginia



http://chronic-wasting-disease.blogspot.com/2009/05/seven-deer-test-positive-for-chronic.html




Terry S. Singeltary Sr.
P.O. Box 42
Bacliff, Texas USA 77518

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