Volume 19, Number 7—July 2013
Dispatch
Asynchronous Onset of Clinical Disease in BSE-Infected Macaques
Judith Montag1, Walter Schulz-Schaeffer, Annette Schrod, Gerhard Hunsmann,
and Dirk Motzkus Author affiliations: German Primate Center, Göttingen, Germany
(J. Montag, A. Schrod, G. Hunsmann, D. Motzkus); University of Göttingen,
Göttingen (W. Schulz-Schaeffer)
Abstract
To estimate the effect of the variability of prion disease onset on primary
bovine spongiform encephalopathy transmission to humans, we studied 6 cynomolgus
macaques. The preclinical incubation period was significantly prolonged in 2
animals, implying that onset of variant Creutzfeldt-Jacob disease in humans
could be more diverse than previously expected.
Prion diseases, such as bovine spongiform encephalopathy (BSE) in cattle,
scrapie in sheep, and Creutzfeldt-Jakob disease (CJD) in humans, are fatal,
transmissible, neurodegenerative disorders associated with the aggregation of an
infectivity-associated isoform (PrPSc) of the cellular prion protein (PrP) (1).
Seventeen years ago, it became apparent that the BSE-infectious agent had
entered the food chain and was identified as the causative agent for a new
variant CJD (vCJD) (2). Since then, several risk assessment studies have
investigated the number of expected vCJD cases in human populations (reviewed in
[3]). Although thousands to millions of consumers of beef products were
estimated to be affected, thus far only a few more than 200 vCJD cases have been
observed worldwide.
This discrepancy was assumed to be attributable to the so-called species
barrier, defined as the hindrance of an infectious agent to change its natural
host. Upon crossing the species barrier, prion diseases often show a low attack
rate in conjunction with a high variability in the preclinical incubation time.
Thus, the consumption of BSE-contaminated products may have led either to a
restricted infection or to a prolonged asymptomatic phase in some exposed
persons. Therefore, concerns have been raised that asymptomatic carriers of vCJD
might exist, posing a risk for unintentional human-to-human transmission.
First indications that transmission of BSE to primates may lead to
variances in the preclinical incubation times were obtained by inoculating
cynomolgus macaques with cattle-derived BSE material (4–6), even though in those
studies not more than 3 animals were used. We have now used a group of 6
macaques that were infected with BSE at a comparable age and kept under
identical and controlled experimental conditions.
The Study Six captive-bred female cynomolgus macaques (Macaca fascicularis,
purchased from the Centre de Recherche en Primatologie, Mauritius) were
inoculated intracerebrally with 1 dose of 50 mg brain homogenate (10% wt/vol)
derived from 11 BSE-infected cattle. Animal experimentation was performed in
accordance with section 8 of the German Animal Protection Law in compliance with
Directive 86/609/EEC. Macaques were housed in a social group, and behavioral
changes were assessed on a daily basis by experienced animal care takers.
After inoculation, all 6 macaques remained healthy and asymptomatic for
>30 months (Table). At 931 days postinfection, 1 animal showed indications of
slight coordination disorders. Within a few days, afferent ataxia developed, and
when the animal was separated from the others animals, she apparently became
tame. After 2 weeks, the animal showed severe dysmetria of the extremities
without obvious myoclonia. Dementia was apparent but could not be diagnosed by
objective measures. For ethical reasons, the animal was euthanized 17 days after
disease onset. Within the next 14 weeks, 3 more animals became symptomatic.
After appearance of neurologic symptoms (ataxia, tremors), the affected animals
were occasionally separated from the group when symptoms became more severe or
attacks from asymptomatic animals occurred. The disease course in these animals
was comparable to that of the first animal, but the progression was slower
(91–103 days).
Figure 1
Figure 1. . Survival of intracerebrally BSE-infected cynomolgus macaques.
Six age- and sex-matched cynomolgus macaques were inoculated intracerebrally
with 50 mg brain homogenate (10% in sucrose) derived from 11 BSE-infected
cattle. Macaques were...
Two of the 6 animals remained asymptomatic for ≈1 additional year. Although
daily monitoring was facilitated by the fact that only 2 macaques remained and
that the caretakers were more experienced to recognize minor changes in
behavior, symptoms were first detected 1,340 and 1,398 days postinfection,
respectively. Clinical signs were similar to those observed in the previous 4
animals. The symptomatic periods before euthanasia for these macaques lasted 103
and 143 days, respectively (Table). Direct comparison revealed that the
difference between the short (931–1,025 days) and the long (1,340–1,398 days)
preclinical incubation time was statistically significant (Figure 1, log-rank
[Mantel-Cox] test, p<0 .05="" div="">
Test results of brain samples from all animals were positive for
macaque-adapted BSE by Western blot analysis. In brief, brain tissue from each
animal was homogenized and subjected to proteinase K (PK) treatment for 1 h at
37°C. Samples were separated on acrylamide gels and transferred to
nitrocellulose membranes. Macaque-adapted BSE (PrPSc) was detected by using the
monoclonal anti-PrP antibody 11C6. PK-resistant PrP was detected in all 6
macaques, confirming that BSE was transmitted to the animals.
Figure 2
Figure 2. . PrPSc profile of macaque-adapted BSE in comparison to human
CJD. Brain homogenates from human sCJD type 1, sCJD type 2, vCJD, and
BSE-infected macaques were subjected to PK treatment, separated...
The individual glycopattern and band migration of macaque-adapted PrPSc was
compared with human sporadic CJD (sCJD) type 1, sCJD type 2, and vCJD.
PK-resistant PrP from BSE-infected macaques co-migrated with type 2 sCJD and was
clearly distinguishable from type 1 sCJD (Figure 2). The glycosylation pattern
of macaque-adapted BSE was comparable with vCJD (6,7), which is characterized by
an overrepresentation of diglycosylated PrPSc (8,9). Using 11C6 antibody (10),
we detected a slightly decreased signal of the diglycosylated PrPSc isoform for
sCJD, vCJD, and macaque-adapted BSE. We assume that this effect is related to a
reduced affinity of the diglycosylated isoform to 11C6 that otherwise shows high
sensitivity to macaque-adapted PrPSc. Nevertheless, direct comparison showed a
higher amount of the diglysosylated PrPSc isoform in vCJD and macaque-adapted
BSE than sCJD, which was also shown with a different monoclonal antibody, 3F4.
This finding confirms that BSE transmission to macaques is comparable with, and
can be used as a model for, human vCJD infection.
Conclusions
Several susceptibility studies using nonhuman primates as a model for human
prion diseases hint to heterogeneity of the preclinical incubation period upon
crossing the species barrier (5,11,12). However, because of the low number of no
more than 3 animals, this variability was not always evident (4). Therefore,
there was an urgent need to determine whether the transmission of BSE to humans
is likely to lead to a similar diversity.
Our study using 6 cynomolgus macaques shows that the transmission of BSE to
primates led to a significantly prolonged asymptomatic phase in 2 animals.
Disease onset is influenced by several factors (13). Our study design enabled us
to exclude that the route of transmission influenced the disease progression
because the infectious agent was injected into the same brain region of each
animal. Also, a limited infectious dose cannot be responsible, as shown by the
attack rate of 100%. In addition, endogenous factors, such as age, the MM
genotype at codon 129 (Table), and housing conditions, were comparable for all
macaques.
Thus, we conclude that the variable asymptomatic phase is most likely
influenced by the infectious agent (14) or the genomic diversity of the macaques
(13). The animals in our study were not inbred. Therefore, differences in the
genomic background may have influenced the time of disease onset. In contrast,
the PrPSc migration patterns of the animals give no indications for different
types or strains that evolved from the mixed BSE inoculum. However, further
studies will have to verify this.
Nevertheless, during the BSE epidemics, the human population with its
natural genomic diversity was also exposed to a nonhomogenous prion source.
Therefore, our study closely mimics the human situation. Our results imply that
a prolonged asymptomatic phase can be expected for vCJD. In light of the
transmissibility of vCJD through blood transfusions (15), our findings emphasize
the need for continued attention to the risks of secondary human-to-human
transmission.
Dr Montag is a microbiologist at the Department of Molecular and Cell
Physiology at the Hannover Medical School. Her primary research interests are
the molecular mechanisms of disease pathology, including prion disorders and
inherited cardiac diseases.
Acknowledgments We thank J.P. Deslys for providing vCJD material.
This study was supported by European Union grant QLK1-CT-2002-01096 and
BMH4-CT-98-6029.
References
snip...see full text ;
Monday, October 10, 2011
EFSA Journal 2011 The European Response to BSE: A Success Story
snip...
EFSA and the European Centre for Disease Prevention and Control (ECDC)
recently delivered a scientific opinion on any possible epidemiological or
molecular association between TSEs in animals and humans (EFSA Panel on
Biological Hazards (BIOHAZ) and ECDC, 2011). This opinion confirmed Classical
BSE prions as the only TSE agents demonstrated to be zoonotic so far but the
possibility that a small proportion of human cases so far classified as
"sporadic" CJD are of zoonotic origin could not be excluded. Moreover,
transmission experiments to non-human primates suggest that some TSE agents in
addition to Classical BSE prions in cattle (namely L-type Atypical BSE,
Classical BSE in sheep, transmissible mink encephalopathy (TME) and chronic
wasting disease (CWD) agents) might have zoonotic potential.
snip...
Thursday, August 12, 2010
Seven main threats for the future linked to prions
First threat
The TSE road map defining the evolution of European policy for protection
against prion diseases is based on a certain numbers of hypotheses some of which
may turn out to be erroneous. In particular, a form of BSE (called atypical
Bovine Spongiform Encephalopathy), recently identified by systematic testing in
aged cattle without clinical signs, may be the origin of classical BSE and thus
potentially constitute a reservoir, which may be impossible to eradicate if a
sporadic origin is confirmed.
***Also, a link is suspected between atypical BSE and some apparently
sporadic cases of Creutzfeldt-Jakob disease in humans. These atypical BSE cases
constitute an unforeseen first threat that could sharply modify the European
approach to prion diseases.
Second threat
snip...
Tuesday, March 05, 2013
A closer look at prion strains Characterization and important implications
Prion
7:2, 99–108; March/April 2013; © 2013 Landes Bioscience
Friday, April 19, 2013
APHIS 2013 Stakeholder Meeting (March 2013) BSE TSE PRION
Monday, May 6, 2013
Warning of mad cow disease threat to blood transfusions
Sunday, May 19, 2013
CJD BLOOD SCREENING, DONORS, AND SILENT CARRIERS House of Commons Written
Answers 16 May 2013
Tuesday, May 21, 2013
CJD, TSE, PRION, BLOOD Abstracts of the 23rd Regional Congress of the
International Society of Blood Transfusion, Amsterdam, The Netherlands, June
2-5, 2013
Sunday, February 10, 2013
Creutzfeldt-Jakob disease (CJD) biannual update (February 2013) Infection
report/CJD
Saturday, December 15, 2012
Bovine spongiform encephalopathy: the effect of oral exposure dose on
attack rate and incubation period in cattle -- an update 5 December 2012
Thursday, February 14, 2013
The Many Faces of Mad Cow Disease Bovine Spongiform Encephalopathy BSE and
TSE prion disease
Tuesday, March 5, 2013
Use of Materials Derived From Cattle in Human Food and Cosmetics; Reopening
of the Comment Period FDA-2004-N-0188-0051 (TSS SUBMISSION)
FDA believes current regulation protects the public from BSE but reopens
comment period due to new studies
Tuesday, November 02, 2010
BSE - ATYPICAL LESION DISTRIBUTION (RBSE 92-21367) statutory (obex only)
diagnostic criteria CVL 1992
Wednesday, April 24, 2013
Chimpanzees Released After 30 Years Of Testing, Brace Yourself For Smiles
TSS
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