Research Article 
Transgenic Mouse Bioassay: Evidence That Rabbits Are Susceptible to a 
Variety of Prion Isolates 
Enric Vidal , Natalia Fernández-Borges , Belén Pintado, Hasier Eraña, 
Montserrat Ordóñez, Mercedes Márquez, Francesca Chianini, Dolors Fondevila, 
Manuel A. Sánchez-Martín, Olivier Andreoletti, Mark P. Dagleish, Martí Pumarola, 
Joaquín Castilla 
PLOS
Published: August 6, 2015 •DOI: 10.1371/journal.ppat.1004977 
Abstract
Interspecies transmission of prions is a well-established phenomenon, both 
experimentally and under field conditions. Upon passage through new hosts, prion 
strains have proven their capacity to change their properties and this is a 
source of strain diversity which needs to be considered when assessing the 
potential risks associated with consumption of prion contaminated protein 
sources. Rabbits were considered for decades to be a prion resistant species 
until proven otherwise recently. To determine the extent of rabbit 
susceptibility to prions and to assess the effects of passage of different prion 
strains through this species a transgenic mouse model overexpressing rabbit PrPC 
was developed (TgRab). Intracerebral challenges with prion strains originating 
from a variety of species including field isolates (ovine SSBP/1 scrapie, Nor98- 
scrapie; cattle BSE, BSE-L and cervid CWD), experimental murine strains (ME7 and 
RML) and experimentally obtained ruminant (sheepBSE) and rabbit (de novo NZW) 
strains were performed. On first passage TgRab were susceptible to the majority 
of prions (Cattle BSE, SheepBSE, BSE-L, de novo NZW, ME7 and RML) tested with 
the exception of SSBP/1 scrapie, CWD and Nor98 scrapie. Furthermore, TgRab were 
capable of propagating strain-specific features such as differences in 
incubation periods, histological brain lesions, abnormal prion (PrPd) deposition 
profiles and proteinase-K (PK) resistant western blotting band patterns. Our 
results confirm previous studies proving that rabbits are not resistant to prion 
infection and show for the first time that rabbits are susceptible to PrPd 
originating in a number of other species. This should be taken into account when 
choosing protein sources to feed rabbits.
Author Summary
Prions, the infectious agents responsible for causing mad cow disease, 
amongst other diseases, can transmit from one species to another. For example, 
Bovine Spongiform Encephalopathy can transmit to humans resulting in invariably 
fatal variant Creutzfeldt-Jakob Disease. We wanted to study the susceptibility 
of rabbits as, until recently, they were considered a prion resistant species. 
Once proven otherwise, we wanted to know which particular prions rabbits were 
susceptible to. With this aim, a transgenic mouse was designed expressing the 
rabbit prion protein gene instead of the corresponding mouse gene to model the 
transmission barrier between rabbits and other species. The resultant mice where 
challenged with several field prion isolates including classical and atypical 
strains of Bovine Spongiform Encephalopathy, sheep Scrapie and cervid Chronic 
Wasting disease. The transgenic mice were susceptible to classical and atypical 
Bovine Spongiform Encephalopathy prions and also to mouse-adapted Scrapie 
prions. This information must be taken into account when assessing the risk of 
using ruminant derived protein as a protein source to feed rabbits. 
snip...
Discussion
This is the first report of in vivo evidence suggesting that TgRab mice are 
susceptible to cross species transmission of prion strains. This not only 
reinforces that rabbits can no longer be considered TSE resistant, but also that 
there is a possibility they could act as a reservoir for other prion strains. As 
such, rabbits must be taken into account when determining the epidemiology of 
several TSE both in relation to the species of origin, especially sympatric 
ones, but also to potential zoonotic transmission.
In previous studies we demonstrated that rabbits were able to propagate 
abnormal prions and that these were transmissible to other rabbits. However, 
this was only one prion strain which was generated de novo in an in vitro PMCA 
assay in rabbit brain homogenate (a spontaneous rabbit prion strain) and on 
first passage it had only a very limited attack rate [23]. This new mouse model, 
which responded in a comparable manner to rabbits when challenged with the same 
in vitro generated rabbit derived inoculum, has allowed us to evaluate the 
TgRab’s susceptibility to a number of actual field prions strains from a variety 
of different species. Although the use of rabbits would have been the most 
appropriate model there are strong, particularly budgetary, limitations due to 
the longer lifespan of rabbits and the need to use level 3 biosafety facilities. 
Thus, a transgenic mouse model overexpressing rabbit PrPC was designed to 
overcome these limitations and allow us to determine its susceptibility to 
different prion strains.
No polymorphisms have been described in the PRNP rabbit gene, therefore 
several mouse transgenic lines were generated expressing rabbit PrPC at 
different expression levels. The line with the highest possible PrPC expression 
levels was selected to allow for easier prion propagation capacity but the 
overexpression was not so high as to generate a spontaneous phenotype at an 
early age which would preclude the attainment of infectivity/susceptibility 
data. The hemizygous TgRab line met these criteria with levels of PrPC between 5 
to 6 times those present in rabbits. The use of transgenic mice overexpressing 
ovine PrPC to obtain the infectivity titer of specific prion isolates has been 
shown to be equivalent to titrations obtained through bioassay in the natural 
host [50]. Phenotyping of the newly developed prion transgenic model was 
essential, especially as the levels of PrPc expression present have not been 
shown to be problematic in other models [41,46]. Eighty percent of the TgRab 
mice presented with a late onset spontaneous neurological disease phenotype (S3 
Fig and S4 Fig) which, fortunately, did not interfere in the interpretation of 
prion susceptibility results. This allowed us to work with a model that 
faithfully reproduced the behavior in rabbits with respect to their capability 
to propagate different prion strains. One cannot exclude the possibility that 
the presence of spontaneous disease might create a toxic environment in the 
brain which artificially enhances the transmission of certain strains. Therefore 
a thorough knowledge of the PrPC overexpression-related changes in uninfected 
controls was essential to identify the true prion disease status and validity of 
susceptibility.
Lesion morphology and profiling within the brain and identification of 
specific PrPd deposition-types allowed unequivocal identification of infected 
animals (either spontaneous or as a result of an inoculation). Further 
biochemical detection of the presence of PrPres by western blotting confirmed 
the ability of morphological techniques to identify an infected animal. 
Additionally, as PrPC overexpression may mask an incipient infection, second 
passages are required to confirm if rabbits are totally resistant to those prion 
isolates to which they initially appeared to be, such as SSBP/1, atypical 
scrapie or CWD, and these experiments are ongoing.
Once validated the TgRab model was used to evaluate which TSE strains were 
able to infect the model (Table 2). Previous attempts in rabbits had concluded 
they were resistant, probably due to incomplete studies and the strong barrier 
of rabbits to propagate prions [34]. Initially classical cattle BSE, the most 
relevant field strain, was tested and found to be infectious on first passage 
with a low attack rate (4/9) and relatively long incubation period (551dpi±10). 
The strain properties observed in the infected TgRab mice (western blotting, 
brain lesion and PrPd deposition profiles) were typical of BSE and 
indistinguishable from those obtained in other BSE murine models [36]. Parallel 
bioassay studies were conducted with the BSE isolate previously amplified in 
vitro using rabbit normal brain homogenate as a substrate (BSE-RaPrPres, this 
inoculum was characterised previously in a TgBov mouse model by our group [36]). 
These animals showed a 100% (12/12) attack rate and a shortened incubation 
period (396dpi ±12 vs 551dpi ±10) compared to the cattle BSE inoculated TgRab 
mice. This reduction already indicated that a transmission barrier between 
species had been overcome thanks to the in vitro adaptation of the cattle BSE-C 
to rabbit PrPC, a second passage was performed from that isolate which also 
showed a 100% attack rate (3/3). Its incubation period was reduced to 322dpi ±12 
(mean ± s.e.m.) indicating further host adaptation (S5 Fig).
SheepBSE, derived from BSE-C, infected TgRab mice with a 100% attack rate 
(9/9), a relatively short incubation time (368±10 dpi) and with lesion and PrPd 
brain profiles identical to those of BSE-C inoculated mice, suggesting that the 
same strain was being propagated through both isolates. This enhanced virulence 
of sheepBSE compared to BSE-C has been previously demonstrated in other 
experimental scenarios [29,51]. The results obtained with sheep scrapie differed 
completely as, in agreement with early experiments in rabbits [34], none of the 
TgRab mice inoculated with SSBP/1 showed any evidence of a prion disease on 
first passage. However, this result does not preclude that, if further in vivo 
SSBP/1 passages were to be performed, the transmission barrier would be crossed. 
As in the case of BSE in the bank vole (Myodes glareolus), where after an 
initial resistance a bank vole adapted BSE strain was obtained which was highly 
transmissible [52,53]. Conversely, ME7 and RML scrapie, both murine adapted 
sheep scrapie strains, infected TgRab mice on first passage and their incubation 
times, PrPres biochemical profiles, lesion profiles and PrPd deposition patterns 
were clearly distinguishable from cattle derived strains. Together these data 
are the first evidence that TgRab mice are not only able to propagate prions but 
they do it maintaining clearly the different distinguishing strain features 
(Figs 1, 3 and 4) which strongly suggests that rabbits may also.
It is noteworthy that both ME7 and RML, which originated from serial 
passages of SSBP/1 in different rodents [54,55], directly propagated in TgRab 
mice on first passage. Conversely, SSBP/1 did not infect TgRab mice on first 
passage. The murine adapted prion strains behaved differently to their parent 
strain, possibly because passage through rodents had selected for a strain 
capable of crossing the rodent species barriers. The situation is analogous to 
CWD which will infect hamsters after initial passage through ferrets [9]. In the 
present work, previous adaptation of scrapie to rodents, possibly resulting in a 
higher sequence identity in some specific and crucial PrP regions with rabbits 
compared to sheep, allowed rodent adapted scrapie prions to misfold rabbit PrPC. 
In previous studies ME7 did not infect rabbits after 4–5 years of incubation, 
with the exception of a single inconclusive case [23,34]. This result is 
difficult to extrapolate since we are discussing different species, of differing 
lifespans and with a species barrier between them. The PrPC overexpression in 
TgRab may have allowed ME7 to propagate more efficiently than in rabbits which 
suggests that if the original rabbit experiments had been performed over the 
maximum lifespan of rabbits ME7 may have propagated on first passage also.
Once BSE in cattle has been virtually controlled, CWD in cervids is the 
animal prion disease with the most repercussions, at least in the North American 
continent. The uncertainty of its transmissibility to humans [56] and its unique 
ability to spread through the free ranging cervid population make its study 
highly relevant with respect to transmissibility to other species. Moreover CWD 
prions are known to be shed and are highly persistent in the environment. 
Rabbits are a sympatric species with cervids. Even though CWD has been shown to 
transmit on first passage to many species including sheep, cattle [57], squirrel 
monkeys [58], cats [59], hamsters [60], ferrets [9], mink [61], bank voles and 
deer mice (Genus Peromyscus) [62] its transmissibility efficiency is relatively 
low with very long incubation periods and low attack rates. For instance, wild 
type mice could not be readily infected, so tga20 mice overexpressing murine 
PrPC were required to prove susceptibility to CWD [63] or required a second 
passage [64]. Another example is the transmission of CWD to cats, which required 
an incubation period of longer than 4 years [59]. The present study showed CWD 
was not able to infect TgRab on first passage (0/12). Further experiments are 
required to confirm the resistance of rabbits to CWD including a blind second 
passage and bioassays with CWD previously passaged in other species, especially 
rodents [9]. This will rule out an analogous situation as the one observed in 
this paper with sheep scrapie whereby SSBP/1 does not transmit to TgRab but 
murine passaged counterparts, ME7 and RML, do.
With respect to the atypical prion strains of purported spontaneous origin 
[18,65,66], BSE-L infected TgRab mice on first passage and, although the attack 
rate was low (3/11), they had the shortest incubation period observed in this 
model so far (221dpi for the first animal to die, mean 280±26dpi). The lesion 
and PrPd deposition brain profiles differed considerably from those of BSE-C. 
None of the TgRab mice inoculated with atypical scrapie showed evidence of a TSE 
with the exception of one animal, euthanized at 742 dpi which, even though no 
histological lesions nor PrPd deposits were present suggestive of infection, it 
was positive by PrPd ELISA. This result could not be confirmed by western 
blotting. However, this ELISA detects PrPd through its affinity to an anionic 
ligand not due to its resistance to protease K so we cannot rule out this single 
mouse was positive. A second passage is ongoing which will determine the 
result.
Initial in vitro experiments predicted that BSE as well as SSBP/1 and CWD 
isolates were able to missfold rabbit PrPC. However, a discrepancy was found 
with the bioassay results since neither SSBP/1 nor CWD infected TgRab mice on 
first passage. Several saPMCA rounds were needed in order to amplify the 
different isolates, varying in number depending of each strain. Thus, it is not 
surprising that on first passage some of the isolates do not transmit.
Besides the PRNP sequence, another component of the transmission barrier is 
the genetic background in which each PrPC is contained. This has been 
demonstrated by infectivity studies showing BSE propagated more efficiently in 
RIII mice than C57/Black mice, two mice strains of the same species with the 
same PRNP gene [67]. Or when the genetic background (i.e. passage through 
different inbred mouse lines) determined not only the incubation period but also 
the propagation of two biochemically different BSE-derived strains [68]. For 
these reasons the belief that rabbits were resistant to prion infection was not 
only attributed to the rabbit PrPC sequence but also to its genetic background. 
To study whether the genetic background of rabbits was responsible for the 
apparent prion resistance, Houdebine’s group generated transgenic rabbits 
expressing an ovine PrPC which was known to easily misfold. Upon inoculation 
with ovine prion strains these rabbits succumbed to prion disease further 
proving that rabbits are not resistant to prions (results published paired with 
this article) and that the genetic background is not a limiting factor 
[37].
The differential susceptibility observed between actual rabbits and the 
transgenic model presented here can be explained by the higher PrPC expression 
levels of TgRab mice. Lower expression mouse lines would probably only be 
susceptible on first passage to strains previously adapted to rabbit PrPC as 
occurs with rabbits. It has taken more than three decades to finally dismiss the 
rabbit as a prion resistant species. We believe that the studies presented here 
confirm that in vitro studies are of great help in interpreting in vivo results, 
leave no room for misinterpretation, and that it can be ascertained that 
rabbits, and probably all other mammal species [21], are susceptible to 
infection by specific prion strains. The prion strain and its species of origin 
determine the extent of susceptibility, but neither rabbit PRNP nor their 
genetic background suggest they are resistant to prion propagation. 
Unfortunately, as with other mammals, the exact molecular mechanisms governing 
the capricious choice of strains that can be propagated in a certain species is 
still unknown.
In light of our results, especially susceptibility to spontaneous cattle 
prions (BSE-L), the restrictions on rabbits being fed ruminant protein should be 
maintained sine die to minimize the chances of any prion strain transmitting to 
rabbits. 
Supporting Information
Figures 
Fig 2 Fig 3 Fig 4 Fig 1 Table 1 Table 2 Fig 2 Fig 3 Fig 4 Fig 1 Table 1 
Table 2 
Citation: Vidal E, Fernández-Borges N, Pintado B, Eraña H, Ordóñez M, 
Márquez M, et al. (2015) Transgenic Mouse Bioassay: Evidence That Rabbits Are 
Susceptible to a Variety of Prion Isolates. PLoS Pathog 11(8): e1004977. 
doi:10.1371/journal.ppat.1004977
Editor: Surachai Supattapone, Dartmouth Medical School, USA, UNITED 
STATES
Received: February 11, 2015; Accepted: May 26, 2015; Published: August 6, 
2015
Copyright: © 2015 Vidal 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
Data Availability: All relevant data are within the paper and its 
Supporting Information files.
Funding: This work was financially supported by 3 national grants from 
Spain [AGL2009-11553-C02-01 (JC), AGL2012-37988-C04-01 (JC) and 
AGL2008-05296-C02 (EV)], a Basque government grant (PI2010-18) (JC), two CTP 
grants (CTP11-P04 and CTP2013-P05) (JC), 3 InterReg grants [EFA205/11 and 
EFA218/11) (JC); EFA282/13—Transprion (MP, DF, EV)], Etortek Research Programs 
2011/2013 (JC) and by Agència de Salut pública de Catalunya, Departament de 
Salut, Generalitat de Catalunya (EV). The funders had no role in study design, 
data collection and analysis, decision to publish, or preparation of the 
manuscript.
Competing interests: The authors have declared that no competing interests 
exist. 
I remember ;
> Despite rabbits no longer being able to be classified as resistant to 
TSEs, an outbreak of “mad rabbit disease” is unlikely. 
Rabbits are not resistant to prion infection 
Francesca Chianinia,1, Natalia Fernández-Borgesb,c,1, Enric Vidald, Louise Gibbarda, Belén Pintadoe, Jorge de Castroc, Suzette A. Priolaf, Scott Hamiltona, Samantha L. Eatona, Jeanie Finlaysona, Yvonne Panga, Philip Steelea, Hugh W. Reida, Mark P. Dagleisha, and Joaquín Castillab,c,g,2 Author Affiliations
Francesca Chianinia,1, Natalia Fernández-Borgesb,c,1, Enric Vidald, Louise Gibbarda, Belén Pintadoe, Jorge de Castroc, Suzette A. Priolaf, Scott Hamiltona, Samantha L. Eatona, Jeanie Finlaysona, Yvonne Panga, Philip Steelea, Hugh W. Reida, Mark P. Dagleisha, and Joaquín Castillab,c,g,2 Author Affiliations
aMoredun Research Institute, Penicuik, Near Edinburgh EH26 0PZ, Scotland, 
United Kingdom; bCIC bioGUNE, Derio 48160, Bizkaia, Spain; gIKERBASQUE, Basque 
Foundation for Science, Bilbao 48011, Bizkaia, Spain; cDepartment of 
Infectology, Scripps Florida, Jupiter, FL 33458; fLaboratory of Persistent Viral 
Diseases, Rocky Mountain Laboratories, National Institute of Allergy and 
Infectious Diseases, National Institutes of Health, Hamilton, MT 59840; dCentre 
de Recerca en Sanitat Animal (CReSA), UAB-IRTA, Universitat Autònoma de 
Barcelona, 08193 Bellaterra, Barcelona, Spain; and eCentro Nacional de 
Biotecnología (CNB), 28049 Cantoblanco, Madrid, Spain Edited by Reed B. Wickner, 
National Institutes of Health, Bethesda, MD, and approved February 16, 2012 
(received for review December 6, 2011) 
Abstract Full Text Authors & Info Figures SI Metrics Related Content 
PDF PDF + SI Abstract The ability of prions to infect some species and not 
others is determined by the transmission barrier. This unexplained phenomenon 
has led to the belief that certain species were not susceptible to transmissible 
spongiform encephalopathies (TSEs) and therefore represented negligible risk to 
human health if consumed. Using the protein misfolding cyclic amplification 
(PMCA) technique, we were able to overcome the species barrier in rabbits, which 
have been classified as TSE resistant for four decades. Rabbit brain homogenate, 
either unseeded or seeded in vitro with disease-related prions obtained from 
different species, was subjected to serial rounds of PMCA. De novo rabbit prions 
produced in vitro from unseeded material were tested for infectivity in rabbits, 
with one of three intracerebrally challenged animals succumbing to disease at 
766 d and displaying all of the characteristics of a TSE, thereby demonstrating 
that leporids are not resistant to prion infection. Material from the brain of 
the clinically affected rabbit containing abnormal prion protein resulted in a 
100% attack rate after its inoculation in transgenic mice overexpressing rabbit 
PrP. Transmissibility to rabbits (>470 d) has been confirmed in 2 of 10 
rabbits after intracerebral challenge. Despite rabbits no longer being able to 
be classified as resistant to TSEs, an outbreak of “mad rabbit disease” is 
unlikely. 
in vitro replication scrapie transmissible spongiform encephalopathy 
Footnotes ↵1F.C. and N.F.-B. contributed equally to this work. 
↵2To whom correspondence should be addressed. E-mail: 
castilla@joaquincastilla.com Author contributions: F.C., N.F.-B., S.A.P., and 
J.d.C. designed research; F.C., N.F.-B., E.V., L.G., B.P., J.d.C., S.A.P., S.H., 
S.L.E., J.F., Y.P., P.S., H.W.R., M.P.D., and J.C. performed research; F.C., 
N.F.-B., E.V., S.A.P., and J.C. contributed new reagents/analytic tools; F.C., 
N.F.-B., E.V., S.A.P., and J.C. analyzed data; and F.C., N.F.-B., S.A.P., 
H.W.R., M.P.D., and J.C. wrote the paper. 
The authors declare no conflict of interest.
This article is a PNAS Direct Submission.
This article contains supporting information online at 
www.pnas.org/lookup/suppl/doi:10.1073/pnas.1120076109/-/DCSupplemental. 
 >>> Despite rabbits no longer being able to be classified as 
resistant to TSEs, an outbreak of “mad rabbit disease” is unlikely. 
seems to be more concern with the latest paper. we went from ‘’Despite 
rabbits no longer being able to be classified as resistant to TSEs, an outbreak 
of “mad rabbit disease” is unlikely’’, in 2012, to ‘’This information must be 
taken into account when assessing the risk of using ruminant derived protein as 
a protein source to feed rabbits.’’ ‘’In light of our results, especially 
susceptibility to spontaneous cattle prions (BSE-L), the restrictions on rabbits 
being fed ruminant protein should be maintained sine die to minimize the chances 
of any prion strain transmitting to rabbits.’’ 
never say never with the tse prion aka mad cow type disease, just to 
correct something, atypical L-type BASE BSE has never been proven to be 
spontaneous under natural conditions in the wild. ...just saying...terry 
P.108: Successful oral challenge of adult cattle with classical BSE 
Sandor Dudas1,*, Kristina Santiago-Mateo1, Tammy Pickles1, Catherine 
Graham2, and Stefanie Czub1 1Canadian Food Inspection Agency; NCAD Lethbridge; 
Lethbridge, Alberta, Canada; 2Nova Scotia Department of Agriculture; Pathology 
Laboratory; Truro, Nova Scotia, Canada 
Classical Bovine spongiform encephalopathy (C-type BSE) is a feed- and 
food-borne fatal neurological disease which can be orally transmitted to cattle 
and humans. Due to the presence of contaminated milk replacer, it is generally 
assumed that cattle become infected early in life as calves and then succumb to 
disease as adults. 
Here we challenged three 14 months old cattle per-orally with 100 grams of 
C-type BSE brain to investigate age-related susceptibility or resistance. During 
incubation, the animals were sampled monthly for blood and feces and subjected 
to standardized testing to identify changes related to neurological disease. 
At 53 months post exposure, progressive signs of central nervous system 
disease were observed in these 3 animals, and they were euthanized. Two of the 
C-BSE animals tested strongly positive using standard BSE rapid tests, however 
in 1 C-type challenged animal, Prion 2015 Poster Abstracts S67 PrPsc was not 
detected using rapid tests for BSE. Subsequent testing resulted in the detection 
of pathologic lesion in unusual brain location and PrPsc detection by PMCA only. 
Our study demonstrates susceptibility of adult cattle to oral transmission 
of classical BSE. We are further examining explanations for the unusual disease 
presentation in the third challenged animal. 
======================== 
***Our study demonstrates susceptibility of adult cattle to oral 
transmission of classical BSE. *** 
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. *** 
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...TSS
=============== 
 DISTRIBUTION: Al, CT, DE, FL, GA, IL, IN, IA, KY, ME, MD, MA, MO, MN, MS, 
NH, NJ, NY, NC, OH, OR, PA, RI, TN, VA, WV, and WI. 
QUANTITY: 2,790 tons of ruminant feed products and 14,000 tons of 
non-ruminant feed products. REASON: The animal feed products may contain protein 
derived from mammalian tissues. 
 RECALLS AND FIELD CORRECTIONS: VETMED -- CLASS IIPRODUCT & CODES: 
Animal feed products, packaged in 5, 25, 50, and 55 pound bags, and in bulk, 
intended for both ruminant and non-ruminant animals. The products are as 
follows: Recall # V-195-1 through V-350-1.
RUMINANT FEED PRODUCTS:
RECALL NO. PRODUCT NO. PRODUCT NAME
V-195-1 40150 B. 30% Calf Pellet V-196-1 40250 B. 16% Calf Pellet V-197-1 
40350 B. 16% Calf Ration V-198-1 40450 B. 18% Calf Starter V-199-1 40600 B. 38% 
Dairy Pellet V-200-1 40650 B. 38% Dairy Pellet V-201-1 40750 B. 16% Dairy Feed 
V-202-1 40950 B. 40% Beef Pellet V-203-1 41150 B. 18% Lamb Starter Pellet 
V-204-1 41250 B. 39% Lamb Conc. Pellet V-205-1 41350 B. 14% Lamb & Beef 
Pellet V-206-1 41450 B. 16% Goat Feed V-207-1 42150 B. 32% Expectation Pellet 
V-208-1 42250 B. Llama & Alpaca Pellet V-209-1 42350 B. 32% Calf Grower 
Pellet V-210-1 42650 B. Llama & Alpaca Crums V-211-1 42750 B. 38% Hay 
Booster 2 V-212-1 42850 B. 25% Pasture Booster V-213-1 43100 B. 16% Grower/Dev 
Pellet V-214-1 43150 B. 16% Grower/Dev Pellet V-215-1 43700 WH 32% Calf Gro 
Pellet V-216-1 43750 WH 32% Calf Gro Pellet V-217-1 43850 B. 38% Dairy Mix 
V-218-1 44250 B. 17% Doe Pellet V-219-1 44350 B. 21% Buck Pellet V-220-1 44450 
Legends Ranch Pellet V-221-1 44500 Legends 17% Breeder Pellet V-222-1 1652 B. 
Vitamin E-20 V-223-1 1614 B. Vitamin A-30 V-224-1 44550 Legends 17% Breeder 
Pellet V-225-1 44650 Legends 13.5% Rut Pellet V-226-1 44750 Deer Starter (J) 
V-227-1 44940 Llama Premix (J) FSC V-228-1 45150 Empire 25% Calf Pellet V-229-1 
45450 Berry Llama Pellet V-230-1 45950 50% Beef Conc. (Meal) V-231-1 46250 B. 
12% Sweet Livestock V-232-1 46350 B. 1440 Bovatec Pellet V-233-1 46400 Liberty 
38% Dairy Pellet V-234-1 46450 Liberty 38% Dairy Pellet V-235-1 47150 B. 14% 
Gold-n-Grower V-236-1 47250 B. 12% Gold-n-Conditioner V-237-1 47450 B. 18% 
Gold-n-Lamb V-238-1 47800 Homeworth Dairy Pellet V-239-1 47850 Homeworth Dairy 
Pellet V-240-1 47900 B. 36% Hi Fat Dairy Pellet V-241-1 47950 B. 36% Hi Fat 
Dairy Pellet V-242-1 48550 B. 16% Calf Pellet CA V-243-1 49200 Mastead Dairy 
Base V-244-1 49300 KLEJKA Dairy Base V-245-1 49650 Deer Premix (J) HFB V-246-1 
49750 39% Lamb Premix (J) HFB V-247-1 49850 Lamb Starter Premix (J) HFB V-248-1 
120850 Brood Cow Deluxe Mineral V-249-1 152850 B. A-D-E Mix
NON-RUMINANT FEED PRODUCTS:
V-250-1 10150 B. Miracle Starter V-251-1 10350 B. 21% Broiler Starter 
V-252-1 10450 B. Pullet Grower & Developer V-253-1 10550 B. 18% Layer 
Breeder Pellets V-254-1 10750 B. 20% Gold Std. Laying Crum V-255-1 10950 B. 17% 
Complete Laying Crums V-256-1 11050 B. 16% Prosperity Layer Crums V-257-1 11100 
B. 40% Poultry Concentrate V-258-1 11150 B. 40% Poultry Concentrate V-259-1 
11250 B. 28% Turkey Starter Crums V-260-1 11350 20% Gig "4" Pellets V-261-1 
11450 B. 16% Prosperity Layer Pellets V-262-1 11550 18% Game Bird Breeder 
Pellets V-263-1 11650 B. 19% Ratite Grower Diet V-264-1 11750 B. 23% Ratite 
Breeder Diet V-265-1 12100 B. 40% Poultry Concentrate Crums V-266-1 12550 B. 32% 
Base Poultry Mix V-267-1 13250 B. 28% Turkey Starter V-268-1 13450 B. 20% 
Poultry Grower V-269-1 14325 B. Game Bird Mix - Coarse V-270-1 20150 B. 18% Pig 
Starter Pellets V-271-1 20250 B. 16% Pig Grower Pellets V-272-1 20450 B. 14% 
Porkmaker 100 Pellets V-273-1 20550 B. 40% Gro 'Em Lean V-274-1 21850 B. 27% 
Hi-Fat Swine Base V-275-1 23000 Mt. Hope Hevy Hog V-276-1 30050 12% Pleasure 
Horse - Sweet V-277-1 30150 Alfa + Performer 10 Sweet V-278-1 30250 14% Grass + 
Perf Sweet V-279-1 30450 12% Wrangler - Complete V-280-1 30550 B. 12% Pleasure 
Horse Pellets V-281-1 30650 B. 32% Gro' N Win Pellets V-282-1 30750 12% Wrangler 
Cubes V-283-1 30950 18% Foal Starter V-284-1 31050 B. 14% Alfa + Dev Pellets 
V-285-1 31150 B. Alfa + Performer 10 Pel V-286-1 31200 Grass +Performer 14 Pel 
V-287-1 31250 Grass +Performer 14 Pel V-288-1 31350 12% Mustang V-289-1 31450 
Endurance - 101 Extruded V-290-1 31550 B. Equine Energy - UK V-291-1 31650 B. 
16% Grass + Dev Pellets V-292-1 31750 16% Grass + Dev Cubes V-293-1 31850 16% 
Grass + Dev Sweet V-294-1 31950 B. 11% Alfa Gro 'N Win Pel V-295-1 32050 B. Sho' 
Win Pellets V-296-1 32250 B. Senior Formula V-297-1 32350 Oscar Horse Mix 
V-298-1 32450 B. Ultimate Finish V-299-1 32550 Crossfire Horse Feed V-300-1 
32650 B. Equine 16% Growth V-301-1 32750 B. Reduced Energy Formula V-302-1 32850 
B. Training Formula V-303-1 32950 B. Cadence Formula V-304-1 33150 B. Track 12 
Horse Feed V-305-1 33350 Spears 16% GR + Dev Cubes V-306-1 33400 B. 14% Supreme 
Horse Pellets V-307-1 33450 B. 14% Supreme Horse Pellets V-308-1 33650 B. Race'N 
Win V-309-1 33750 B. 14% Prominent Horse Feed V-310-1 33850 B. Unbeetable Horse 
Feed V-311-1 34750 Cargill Senior Horse V-312-1 34850 Cargill Vitality Gold 
V-313-1 35150 Chagrin 12% Sweet Fd V-314-1 35250 Smith Pure Pleasure V-315-1 
35750 Roundup 10% Horse Pellets V-316-1 35850 12% Summerglo Horse V-317-1 36255 
B. Grass +Min&VitBase - Mexico V-318-1 36850 Miller's 12% Horse Feed V-319-1 
37155 B. Gro'Win Base Mix - Mexico V-320-1 38000 B. 32% Premium Mixer Pellets 
V-321-1 38050 B. 32% Premium Mixer Pellets V-322-1 38100 36% Maintenance Mixer 
Pellets V-323-1 38150 36% Maintenance Mixer Pellets V-324-1 50150 Terramycin 
Crumbles 
=========================
V-325-1 60105 16% Rabbit Pellets V-326-1 60125 16% Rabbit Pellets V-327-1 
60150 B. 16% Rabbit Pellets V-328-1 60205 18% Rabbit Developer V-329-1 60250 B. 
18% Rabbit Developer V-330-1 60450 B. 16% Rabbit Maintenance 
=========================
V-331-1 90150 B. Buckeye Scratch V-332-1 90225 Gold Standard Scratch 
V-333-1 90250 Gold Standard Scratch V-334-1 90350 Intermediate Scratch V-335-1 
90450 B. Chick Grains V-336-1 90525 B. Shelled Corn V-337-1 90550 B. Shelled 
Corn V-338-1 90650 B. Cracked Corn V-339-1 90825 B. Fine Cracked Corn V-340-1 
90850 B. Fine Cracked Corn V-341-1 91000 Steam Flaked Corn V-342-1 91050 Steam 
Flaked Corn V-343-1 91750 Oats - HP Crimped V-344-1 91850 B. HP Sweet Crimped 
Oats V-345-1 95550 Land O' Lakes Shelled Corn V-346-1 95650 Land O' Cracked Corn 
V-347-1 95850 Land O' Lakes Chick Crack V-348-1 100850 B. Alfalfa Pellets 
V-349-1 101850 Cooked Full Fat Soybean V-350-1 122200 Magnatone M-4-B Pels Bulk 
MANUFACTURER: Buckeye Feed Mills, Dalton, Ohio. RECALLED BY: Manufacturer 
visited local customers on April 17, 2001. On April 18 and 19, 2001, 
manufacturer mailed and faxed recall notices. Firm initiated recall is ongoing. 
DISTRIBUTION: Al, CT, DE, FL, GA, IL, IN, IA, KY, ME, MD, MA, MO, MN, MS, 
NH, NJ, NY, NC, OH, OR, PA, RI, TN, VA, WV, and WI. 
QUANTITY: 2,790 tons of ruminant feed products and 14,000 tons of 
non-ruminant feed products. 
REASON: The animal feed products may contain protein derived from mammalian 
tissues. 
 10 years post mad cow feed ban August 1997 
10,000,000+ LBS. of PROHIBITED BANNED MAD COW FEED I.E. BLOOD LACED MBM IN 
COMMERCE USA 2007 
Date: March 21, 2007 at 2:27 pm PST 
RECALLS AND FIELD CORRECTIONS: VETERINARY MEDICINES -- CLASS II 
PRODUCT 
Bulk cattle feed made with recalled Darling's 85% Blood Meal, Flash Dried, 
Recall # V-024-2007 
CODE 
Cattle feed delivered between 01/12/2007 and 01/26/2007 
RECALLING FIRM/MANUFACTURER 
Pfeiffer, Arno, Inc, Greenbush, WI. by conversation on February 5, 2007. 
Firm initiated recall is ongoing. 
REASON 
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. 
VOLUME OF PRODUCT IN COMMERCE 
42,090 lbs. 
DISTRIBUTION 
WI 
___________________________________ 
PRODUCT 
Custom dairy premix products: MNM ALL PURPOSE Pellet, HILLSIDE/CDL Prot- 
Buffer Meal, LEE, M.-CLOSE UP PX Pellet, HIGH DESERT/ GHC LACT Meal, TATARKA, M 
CUST PROT Meal, SUNRIDGE/CDL PROTEIN Blend, LOURENZO, K PVM DAIRY Meal, DOUBLE B 
DAIRY/GHC LAC Mineral, WEST PIONT/GHC CLOSEUP Mineral, WEST POINT/GHC LACT Meal, 
JENKS, J/COMPASS PROTEIN Meal, COPPINI - 8# SPECIAL DAIRY Mix, GULICK, L-LACT 
Meal (Bulk), TRIPLE J - PROTEIN/LACTATION, ROCK CREEK/GHC MILK Mineral, 
BETTENCOURT/GHC S.SIDE MK-MN, BETTENCOURT #1/GHC MILK MINR, V&C DAIRY/GHC 
LACT Meal, VEENSTRA, F/GHC LACT Meal, SMUTNY, A- BYPASS ML W/SMARTA, Recall # 
V-025-2007 
CODE 
The firm does not utilize a code - only shipping documentation with 
commodity and weights identified. 
RECALLING FIRM/MANUFACTURER 
Rangen, Inc, Buhl, ID, by letters on February 13 and 14, 2007. Firm 
initiated recall is complete. 
REASON 
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. 
VOLUME OF PRODUCT IN COMMERCE 
9,997,976 lbs. 
DISTRIBUTION 
ID and NV 
END OF ENFORCEMENT REPORT FOR MARCH 21, 2007 
16 years post mad cow feed ban August 1997
2013
Sunday, December 15, 2013 
FDA PART 589 -- SUBSTANCES PROHIBITED FROM USE IN ANIMAL FOOD OR FEED 
VIOLATIONS OFFICIAL ACTION INDICATED OIA UPDATE DECEMBER 2013 UPDATE
17 years post mad cow feed ban August 1997
Tuesday, December 23, 2014 
FDA PART 589 -- SUBSTANCES PROHIBITED FROM USE IN ANIMAL FOOD OR FEED 
VIOLATIONS OFFICIAL ACTION INDICATED OAI UPDATE DECEMBER 2014 BSE TSE PRION 
Sunday, June 14, 2015 
Larry’s Custom Meats Inc. Recalls Beef Tongue Products That May Contain 
Specified Risk Materials BSE TSE Prion
DR. DEHAVEN: 
snip...
*** 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. 
snip... 
 http://www.usda.gov/wps/portal/!ut/p/_s.7_0_A/7_0_1OB?contentidonly=true&contentid=2004/03/0106.html 
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 worlds 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.
We can’t say it’s not feed related, agreed Dr. Linda Detwiler, an official 
with the USDA during the Clinton Administration now at Mississippi State.
In the May 1 email to me, USDA’s Cole backed off a bit. No one knows the 
origins of atypical cases of BSE, she said
Few scientists would argue that the one California cow which never was 
headed to the U.S. food supply represents a health hazard. 
But many maintain that the current surveillance is insufficient. 
Dr. Kurt Giles, an expert in neurogenerative diseases now at the University 
of California, San Francisco, was at Oxford during the British outbreak. 
He told me USDA’s assurances about safety today remind him of British 
statements during the 1980s. 
It is so reminiscent of that absolute certainty, he said.
Robert Bazell is NBC's chief science and medical correspondent. Follow him 
on Facebook and on Twitter @RobertBazellNBC 
THE USDA JUNE 2004 ENHANCED BSE SURVEILLANCE PROGRAM WAS TERRIBLY FLAWED ; 
CDC DR. PAUL BROWN TSE EXPERT COMMENTS 2006
In an article today for United Press International, science reporter Steve 
Mitchell writes:
Analysis: What that mad cow means
By STEVE MITCHELL UPI Senior Medical Correspondent
WASHINGTON, March 15 (UPI) -- The U.S. Department of Agriculture was quick 
to assure the public earlier this week that the third case of mad cow disease 
did not pose a risk to them, but what federal officials have not acknowledged is 
that this latest case indicates the deadly disease has been circulating in U.S. 
herds for at least a decade.
The second case, which was detected last year in a Texas cow and which USDA 
officials were reluctant to verify, was approximately 12 years old.
These two cases (the latest was detected in an Alabama cow) present a 
picture of the disease having been here for 10 years or so, since it is thought 
that cows usually contract the disease from contaminated feed they consume as 
calves. The concern is that humans can contract a fatal, incurable, 
brain-wasting illness from consuming beef products contaminated with the mad cow 
pathogen.
"The fact the Texas cow showed up fairly clearly implied the existence of 
other undetected cases," Dr. Paul Brown, former medical director of the National 
Institutes of Health's Laboratory for Central Nervous System Studies and an 
expert on mad cow-like diseases, told United Press International. "The question 
was, 'How many?' and we still can't answer that."
Brown, who is preparing a scientific paper based on the latest two mad cow 
cases to estimate the maximum number of infected cows that occurred in the 
United States, said he has "absolutely no confidence in USDA tests before one 
year ago" because of the agency's reluctance to retest the Texas cow that 
initially tested positive.
USDA officials finally retested the cow and confirmed it was infected seven 
months later, but only at the insistence of the agency's inspector 
general.
"Everything they did on the Texas cow makes everything they did before 2005 
suspect," Brown said.
SNIP...
UPI requested detailed records about animals tested under the USDA's 
surveillance plan via the Freedom of Information Act in May 2004 but nearly two 
years later has not received any corresponding documents from the agency, 
despite a federal law requiring agencies to comply within 30 days. This leaves 
open the question of whether the USDA is withholding the information, does not 
have the information or is so haphazardly organized that it cannot locate 
it.
SNIP...
Markus Moser, a molecular biologist and chief executive officer of 
Prionics, a Swiss firm that manufactures BSE test kits, told UPI one concern is 
that if people are infected, the mad cow pathogen could become "humanized" or 
more easily transmitted from person to person.
"Transmission would be much easier, through all kinds of medical 
procedures" and even through the blood supply, Moser said.
© Copyright 2006 United Press International, Inc. All Rights Reserved 
CDC - Bovine Spongiform Encephalopathy and Variant Creutzfeldt ... Dr. Paul 
Brown is Senior Research Scientist in the Laboratory of Central Nervous System 
... Address for correspondence: Paul Brown, Building 36, Room 4A-05, ... 
PAUL BROWN COMMENT TO ME ON THIS ISSUE 
Tuesday, September 12, 2006 11:10 AM
"Actually, Terry, I have been critical of the USDA handling of the mad cow 
issue for some years, and with Linda Detwiler and others sent lengthy detailed 
critiques and recommendations to both the USDA and the Canadian Food Agency." 
........TSS 
FRANCE HAVE AN EPIDEMIC OF SPONTANEOUS ATYPICAL BSE ‘’LOL’’
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. 
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 
spontaneous TSE prion, that's wishful thinking. on the other hand, if 
spontaneous did ever happen (never once documented in the field), it would be 
our worst nightmare, due to feed. just saying. 
*** 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.*** 
>>> 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. 
<<< 
2014 
***Moreover, L-BSE has been transmitted more easily to transgenic mice 
overexpressing a human PrP [13,14] or to primates [15,16] than C-BSE. 
***It has been suggested that some sporadic CJD subtypes in humans may 
result from an exposure to the L-BSE agent. 
*** Lending support to this hypothesis, pathological and biochemical 
similarities have been observed between L-BSE and an sCJD subtype (MV genotype 
at codon 129 of PRNP) [17], and between L-BSE infected non-human primate and 
another sCJD subtype (MM genotype) [15]. 
snip... 
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
Acquired transmissibility of sheep-passaged L-type bovine spongiform 
encephalopathy prion to wild-type mice 
Short report Acquired transmissibility of sheep-passaged L-type bovine 
spongiform encephalopathy prion to wild-type mice Hiroyuki Okada*, Kentaro 
Masujin*, Kohtaro Miyazawa and Takashi Yokoyama 
* Corresponding authors: Hiroyuki Okada okadahi@affrc.go.jp - Kentaro 
Masujin masujin@affrc.go.jp 
Author Affiliations
National Institute of Animal Health, National Agriculture and Food Research 
Organization (NARO), Tsukuba, Ibaraki, Japan 
For all author emails, please log on. 
Veterinary Research 2015, 46:81 doi:10.1186/s13567-015-0211-2
The electronic version of this article is the complete one and can be found 
online at: http://www.veterinaryresearch.org/content/46/1/81 
Received: 3 February 2015 Accepted: 8 June 2015 Published: 13 July 2015 
© 2015 Okada et al. 
The transmission experiment reported here shows that the host range of 
L-BSE prions can be extended by inter-species transmission. Further experimental 
transmission of L-BSE/sheep along with L-BSE into humanized PrP mice will be 
necessary to evaluate the potential risk for humans. 
see full text ; 
Wednesday, July 29, 2015 
Acquired transmissibility of sheep-passaged L-type bovine spongiform 
encephalopathy prion to wild-type mice 
IBNC Tauopathy or TSE Prion disease, it appears, no one is sure
Posted by flounder on 03 Jul 2015 at 16:53 GMT
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?
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. 
SNIP...
Discussion
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. 
***In addition, non-human primates are specifically susceptible for 
atypical BSE as demonstrated by an approximately 50% shortened incubation time 
for L-type BSE as compared to C-type. Considering the current scientific 
information available, it cannot be assumed that these different BSE types pose 
the same human health risks as C-type BSE or that these risks are mitigated by 
the same protective measures. 
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) 
 SUMMARY REPORT CALIFORNIA BOVINE SPONGIFORM ENCEPHALOPATHY CASE 
INVESTIGATION JULY 2012 (ATYPICAL L-TYPE BASE)
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)
snip...
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. 
TEXAS ATYPICAL H-BSE MAD COW CASE
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)) 
ALABAMA ATYPICAL H-TYPE GENETIC BSE 
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 
BSE ATYPICAL TEXAS AND ALABAMA UPDATE JANUARY 20, 2007 
LAST MAD COW IN USA, IN CALIFORNIA, WAS ATYPICAL L-TYPE BASE BSE TSE PRION 
DISEASE Thursday, February 20, 2014 
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 ; 
OR, what the Honorable Phyllis Fong of the OIG found ;
Audit Report Animal and Plant Health Inspection Service Bovine Spongiform 
Encephalopathy (BSE) Surveillance Program  Phase II and Food Safety and 
Inspection Service
Controls Over BSE Sampling, Specified Risk Materials, and Advanced Meat 
Recovery Products - Phase III
Report No. 50601-10-KC January 2006
Finding 2 Inherent Challenges in Identifying and Testing High-Risk Cattle 
Still Remain 
ALSO, PLEASE SEE ; 
31 Jan 2015 at 20:14 GMT 
*** Ruminant feed ban for cervids in the United States? *** 
31 Jan 2015 at 20:14 GMT 
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: maf12@cam.ac.uk Jürgen 
A. Richt College of Veterinary Medicine, Kansas State University, K224B Mosier 
Hall, Manhattan, Kansas 66506-5601, USA NATURE|Vol 457|26 February 2009 
Thursday, July 24, 2014 
*** Protocol for further laboratory investigations into the distribution of 
infectivity of Atypical BSE SCIENTIFIC REPORT OF EFSA New protocol for Atypical 
BSE investigations 
 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 
19 May 2010 at 21:21 GMT 
*** Singeltary reply ; Molecular, Biochemical and Genetic Characteristics 
of BSE in Canada Singeltary reply ; 
*** ATYPICAL BSE AND POTENTIAL FOR ANIMAL PROTEIN FEED TO BE A LINK THERE 
FROM ***
P.9.21 Molecular characterization of BSE in Canada 
Jianmin Yang1, Sandor Dudas2, Catherine Graham2, Markus Czub3, Tim 
McAllister1, Stefanie Czub1 1Agriculture and Agri-Food Canada Research Centre, 
Canada; 2National and OIE BSE Reference Laboratory, Canada; 3University of 
Calgary, Canada 
Background: Three BSE types (classical and two atypical) have been 
identified on the basis of molecular characteristics of the misfolded protein 
associated with the disease. To date, each of these three types have been 
detected in Canadian cattle. Objectives: This study was conducted to further 
characterize the 16 Canadian BSE cases based on the biochemical properties of 
there associated PrPres. 
Methods: Immuno-reactivity, molecular weight, glycoform profiles and 
relative proteinase K sensitivity of the PrPres from each of the 16 confirmed 
Canadian BSE cases was determined using modified Western blot analysis. 
Results: Fourteen of the 16 Canadian BSE cases were C type, 1 was H type 
and 1 was L type. The Canadian H and L-type BSE cases exhibited size shifts and 
changes in glycosylation similar to other atypical BSE cases. PK digestion under 
mild and stringent conditions revealed a reduced protease resistance of the 
atypical cases compared to the C-type cases. N terminal- specific antibodies 
bound to PrPres from H type but not from C or L type. The C-terminal-specific 
antibodies resulted in a shift in the glycoform profile and detected a fourth 
band in the Canadian H-type BSE. 
Discussion: The C, L and H type BSE cases in Canada exhibit molecular 
characteristics similar to those described for classical and atypical BSE cases 
from Europe and Japan. This supports the theory that the importation of BSE 
contaminated feedstuff is the source of C-type BSE in Canada. * It also suggests 
a similar cause or source for atypical BSE in these countries.
*** It also suggests a similar cause or source for atypical BSE in these 
countries. *** 
Discussion: The C, L and H type BSE cases in Canada exhibit molecular 
characteristics similar to those described for classical and atypical BSE cases 
from Europe and Japan. *** This supports the theory that the importation of BSE 
contaminated feedstuff is the source of C-type BSE in Canada. *** It also 
suggests a similar cause or source for atypical BSE in these countries. *** 
see page 176 of 201 pages...tss 
Thursday, July 24, 2014 
*** Protocol for further laboratory investigations into the distribution of 
infectivity of Atypical BSE SCIENTIFIC REPORT OF EFSA New protocol for Atypical 
BSE investigations 
Guidance for Industry Ensuring Safety of Animal Feed Maintained and Fed 
On-Farm Draft Guidance FDA-2014-D-1180 Singeltary Comment
Thursday, February 20, 2014
***Oral Transmission of L-type Bovine Spongiform Encephalopathy in Primate 
Model
***Infectivity in skeletal muscle of BASE-infected cattle
***feedstuffs- It also suggests a similar cause or source for atypical BSE 
in these countries. ***
***Also, a link is suspected between atypical BSE and some apparently 
sporadic cases of Creutzfeldt-Jakob disease in humans.
full text ;
atypical L-type BASE BSE
http://transmissiblespongiformencephalopathy.blogspot.com/2012/04/update-from-usda-regarding-detection-of.html 
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. 
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. 
snip... 
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 ; 
.....snip 
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. 
Wednesday, July 29, 2015 
Further characterisation of transmissible spongiform encephalopathy 
phenotypes after inoculation of cattle with two temporally separated sources of 
sheep scrapie from Great Britain
Wednesday, July 29, 2015 
Progressive accumulation of the abnormal conformer of the prion protein and 
spongiform encephalopathy in the obex of nonsymptomatic and symptomatic Rocky 
Mountain elk (Cervus elaphus nelsoni) with chronic wasting disease
Wednesday, July 29, 2015 
Porcine Prion Protein Amyloid or mad pig disease PSE 
Wednesday, July 29, 2015 
Acquired transmissibility of sheep-passaged L-type bovine spongiform 
encephalopathy prion to wild-type mice 
IBNC Tauopathy or TSE Prion disease, it appears, no one is sure
Posted by flounder on 03 Jul 2015 at 16:53 GMT
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?
2015 FDA UPDATE BSE
Tuesday, August 4, 2015 
*** FDA U.S. Measures to Protect Against BSE ***
BANNED MAD COW FEED IN COMMERCE IN ALABAMA 
______________________________ 
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. 
REASON 
Dairy and poultry feeds were possibly contaminated with ruminant based 
protein. 
VOLUME OF PRODUCT IN COMMERCE 
477.72 tons 
DISTRIBUTION 
AL 
______________________________ 
PRODUCT 
a) Dairy feed, custom, Recall # V-134-6; 
b) Custom Dairy Feed with Monensin, Recall # V-135-6. 
CODE 
None. Bulk product 
RECALLING FIRM/MANUFACTURER 
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. 
REASON 
Possible contamination of dairy feeds with ruminant derived meat and bone 
meal. 
VOLUME OF PRODUCT IN COMMERCE 
1,484 tons 
DISTRIBUTION 
TN and WV 
END OF ENFORCEMENT REPORT FOR SEPTEMBER 6, 2006 
### 
RECALLS AND FIELD CORRECTIONS: VETERINARY MEDICINE - CLASS II 
______________________________ 
PRODUCT 
Bulk custom made dairy feed, Recall # V-115-6 
CODE 
None 
RECALLING FIRM/MANUFACTURER 
Hiseville Feed & Seed Co., Hiseville, KY, by telephone and letter on or 
about July 14, 2006. FDA initiated recall is ongoing. 
REASON 
Custom made feeds contain ingredient called Pro-Lak which may contain 
ruminant derived meat and bone meal. 
VOLUME OF PRODUCT IN COMMERCE 
Approximately 2,223 tons 
DISTRIBUTION 
KY 
______________________________ 
PRODUCT 
Bulk custom made dairy feed, Recall # V-116-6 
CODE 
None 
RECALLING FIRM/MANUFACTURER 
Rips Farm Center, Tollesboro, KY, by telephone and letter on July 14, 2006. 
FDA initiated recall is ongoing. 
REASON 
Custom made feeds contain ingredient called Pro-Lak which may contain 
ruminant derived meat and bone meal. 
VOLUME OF PRODUCT IN COMMERCE 
1,220 tons 
DISTRIBUTION 
KY 
______________________________ 
PRODUCT 
Bulk custom made dairy feed, Recall # V-117-6 
CODE 
None 
RECALLING FIRM/MANUFACTURER 
Kentwood Co-op, Kentwood, LA, by telephone on June 27, 2006. FDA initiated 
recall is completed. 
REASON 
Possible contamination of animal feed ingredients, including ingredients 
that are used in feed for dairy animals, with ruminant derived meat and bone 
meal. 
VOLUME OF PRODUCT IN COMMERCE 
40 tons 
DISTRIBUTION 
LA and MS 
______________________________ 
PRODUCT 
Bulk Dairy Feed, Recall V-118-6 
CODE 
None 
RECALLING FIRM/MANUFACTURER 
Cal Maine Foods, Inc., Edwards, MS, by telephone on June 26, 2006. FDA 
initiated recall is complete. 
REASON 
Possible contamination of animal feed ingredients, including ingredients 
that are used in feed for dairy animals, with ruminant derived meat and bone 
meal. 
VOLUME OF PRODUCT IN COMMERCE 
7,150 tons 
DISTRIBUTION 
MS 
______________________________ 
PRODUCT 
Bulk custom dairy pre-mixes, Recall # V-119-6 
CODE 
None 
RECALLING FIRM/MANUFACTURER 
Walthall County Co-op, Tylertown, MS, by telephone on June 26, 2006. Firm 
initiated recall is complete. 
REASON 
Possible contamination of dairy animal feeds with ruminant derived meat and 
bone meal. 
VOLUME OF PRODUCT IN COMMERCE 
87 tons 
DISTRIBUTION 
MS 
______________________________ 
PRODUCT 
Bulk custom dairy pre-mixes, Recall # V-120-6 
CODE 
None 
RECALLING FIRM/MANUFACTURER 
Ware Milling Inc., Houston, MS, by telephone on June 23, 2006. Firm 
initiated recall is complete. 
REASON 
Possible contamination of dairy animal feeds with ruminant derived meat and 
bone meal. 
VOLUME OF PRODUCT IN COMMERCE 
350 tons 
DISTRIBUTION 
AL and MS 
______________________________ 
PRODUCT 
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 
CODE 
All products manufactured from 02/01/2005 until 06/20/2006 
RECALLING FIRM/MANUFACTURER 
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. 
REASON 
Poultry and fish feeds which were possibly contaminated with ruminant based 
protein were not labeled as "Do not feed to ruminants". 
VOLUME OF PRODUCT IN COMMERCE 
7,541-50 lb bags 
DISTRIBUTION 
AL, GA, MS, and TN 
END OF ENFORCEMENT REPORT FOR AUGUST 9, 2006 
### 
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". 
VOLUME OF PRODUCT IN COMMERCE 125 tons 
DISTRIBUTION AL and FL 
END OF ENFORCEMENT REPORT FOR AUGUST 2, 2006 
### 
MAD COW FEED RECALL USA EQUALS 10,878.06 TONS NATIONWIDE Sun Jul 16, 2006 
09:22 71.248.128.67 
RECALLS AND FIELD CORRECTIONS: VETERINARY MEDICINE -- CLASS II 
______________________________ 
PRODUCT 
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; 
c) PRO-PAK, MARINE & ANIMAL PROTEIN CONCENTRATE FOR USE IN ANIMAL FEED, 
Recall # V-081-6; 
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. 
REASON 
Possible contamination of animal feeds with ruminent derived meat and bone 
meal. 
VOLUME OF PRODUCT IN COMMERCE 10,878.06 tons 
DISTRIBUTION Nationwide 
END OF ENFORCEMENT REPORT FOR July 12, 2006 
### 
what about that ALABAMA MAD COW, AND MAD COW FEED THERE FROM IN THAT STATE 
??? 
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) 
BANNED MAD COW FEED IN COMMERCE IN ALABAMA 
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. 
REASON 
Dairy and poultry feeds were possibly contaminated with ruminant based 
protein.
VOLUME OF PRODUCT IN COMMERCE 477.72 tons 
DISTRIBUTION AL 
______________________________
Saturday, March 21, 2015 
Canada and United States Creutzfeldt Jakob TSE Prion Disease Incidence 
Rates Increasing 
Terry S. Singeltary Sr.
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