- Interspecies Transmission of Chronic Wasting Disease
Prions to Squirrel Monkeys (Saimiri sciureus)
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- Chronic wasting disease (CWD) is an emerging prion disease
of deer and elk. The risk of CWD transmission to humans following exposure
to CWD-infected tissues is unknown. To assess the susceptibility of nonhuman
primates to CWD, two squirrel monkeys were inoculated with brain tissue
from a CWD-infected mule deer. The CWD-inoculated squirrel monkeys developed
a progressive neurodegenerative disease and were euthanized at 31 and
34 months postinfection. Brain tissue from the CWD-infected squirrel monkeys
contained the abnormal isoform of the prion protein, PrP-res, and displayed
spongiform degeneration. This is the first reported transmission of CWD
to primates.
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- Chronic wasting disease (CWD) is a prion disease of elk
and deer in North America that was first identified at cervid research
facilities in Colorado and Wyoming in the late 1960s (17, 18). CWD has
been identified on cervid game farms from Montana to New York and has
been diagnosed in wild deer and elk in Colorado, Wyoming, Nebraska, South
Dakota, Wisconsin, New Mexico, Illinois, and Utah and in Saskatchewan,
Canada (1, 14, 15). The geographic distribution of CWD in deer and elk
has been expanding and will likely result in an increase in human exposure
to the CWD agent. Although there have been no cases of human prion disease
linked to CWD infection, the risk of interspecies transmission of CWD
to humans following consumption of CWD-infected tissues is uncertain
(5, 13).
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- One approach to assess the susceptibility of humans to
animal prion diseases is by experimental transmission to nonhuman primates
(9-11). To investigate the susceptibility of nonhuman primates to CWD,
two adult female squirrel monkeys (Saimiri sciureus) were intracerebrally
(i.c.) inoculated with 200 µl of a 20% (wt/vol) brain homogenate
from a female mule deer in the clinical phase of CWD (inoculum was a gift
from Elizabeth Williams, Department of Veterinary Sciences, University
of Wyoming, Laramie, WY). Both CWD- inoculated squirrel monkeys developed
a progressive neurological disease and were euthanized at the terminal
stages of disease at 31 and 34 months postinfection, respectively (data
on clinical symptoms and the time to onset of disease were not available).
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- To determine whether the abnormal form of the prion protein,
PrP- res, was present in the CWD-infected squirrel monkeys, brain homogenates
were analyzed by Western blotting as previously described using the anti-PrP
monoclonal antibody 6H4 (Prionics AG, Switzerland) (2). For this analysis,
a 5% (wt/vol) brain homogenate in Dulbecco's phosphate-buffered saline
(Mediatech, Inc.) from CWD- infected squirrel monkeys, a CWD-infected elk,
or an uninfected mouse was either digested with proteinase K (PK) (4 U/ml;
United States Biochemical) for 1 h at 37°C with agitation or was not
digested with PK. In the samples that were not digested with PK, PrP
migrated between 21 and 35 kDa in the CWD-infected squirrel monkeys (Fig.
1, lanes 1 and 2) and between 30 and 35 kDa in the CWD-infected elk (Fig.
1, lane 3) and in the uninfected mouse sample (Fig. 1, lane 4). In the
samples that were digested with PK, PrP-res were detected in the two CWD-infected
squirrel monkeys (Fig. 1, lanes 5 and 6) and in the CWD-infected elk sample
(Fig. 1, lane 7). In the PK-digested uninfected mouse brain, PrP was not
detected (Fig. 1, lane 8), indicating that PK digestion completely removed
the PK-sensitive isoform of PrP. In both CWD-infected squirrel monkeys,
the migration of the three PrP-res polypeptides on sodium dodecyl sulfate-polyacrylamide
gels was similar. The diglycosylated PrP-res polypeptide migrated at 30
kDa similar to what has been reported for squirrel monkeys infected with
sporadic Creutzfeldt-Jakob disease (CJD), kuru, and scrapie (4). The
relative abundance of PrP-res in the brain from the squirrel monkey that
was sacrificed at 34 months postinfection (Fig. 1, lane 5) was greater
than that in the squirrel monkey sacrificed at 31 months postinfection
(Fig. 1, lane 6) and may represent differences in the state of disease
progression when the animals were sacrificed.
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- Histological examination of the brain, brain stem, and
spinal cord from the squirrel monkey that was euthanized at 31 months
postinfection revealed widespread spongiform changes that are consistent
with CWD-induced neurodegeneration. Spongiform lesions in the neuropil
were predominantly located in subcortical gray matter structures of the
forebrain. There was widespread spongiform change in the putamen, caudate
nucleus, nucleus accumbens, lateral and medial hypothalamus, hippocampal
formation (CA 1), amygdala, and dorsomedial thalamus (Fig. 2). Diffuse
spongiosis was found in the interpeduncular nucleus and substantia nigra
in the midbrain and in the reticular formation of the pons and medulla.
Due to the limited number of histological sections, a detailed comparison
of the neuropathology in CWD-infected squirrel monkeys and other prion
transmission studies in squirrel monkeys was not possible.
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- The time to terminal disease following inoculation of
squirrel monkeys with the CWD agent, 31 and 34 months, was longer than
for squirrel monkeys that were i.c. inoculated with transmissible mink
encephalopathy agent (9 to 12 months) and scrapie agent (16 months) but
is within the reported range of the time to terminal disease following
i.c. inoculation with sporadic CJD (11 to 37 months) and kuru (10 to 48
months) (6, 8). This variation in disease progression following experimental
transmission of sporadic CJD, kuru, and CWD to squirrel monkeys could
be due to differences in the inoculation dose, strain of the prion agent,
or the ability to establish infection upon interspecies transmission.
Regardless, this study illustrates that a nonhuman primate can develop
a prion disease following i.c. inoculation with a brain homogenate from
a CWD-infected mule deer.
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- Direct comparison of the ability of the CWD agent to
cause disease in squirrel monkeys following experimental i.c. inoculation
and the susceptibility of humans to CWD infection must be interpreted
with caution. Although squirrel monkeys are susceptible to experimental
infection with kuru and CJD, they are also susceptible to experimental
infection with scrapie (8), and there is no epidemiological evidence to
suggest that scrapie can be transmitted to humans (16). These data suggest,
following direct cerebral inoculation, squirrel monkeys may not be a good
experimental model for assessing human susceptibility to animal prion
diseases. Oral exposure is the likely natural route of human exposure
to CWD, and in experimental animals, this route is much less efficient
at causing disease than i.c. inoculation (3, 7, 12). Therefore, the ability
of scrapie and CWD to cause disease in primates by oral infection needs
to be established to further resolve the issue of susceptibility of humans
to CWD infection.
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- Richard Marsh, who performed the experimental transmission
of CWD to squirrel monkeys, died in 1997 before these experiments were
completed. Due to the emergence of CWD in deer and elk and the potential
risk for CWD transmission to humans, we present his findings with additional
tissue analysis.
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- We thank Al Jenny, USDA-APHIS-VS-NVSL for the gift of
the CWD- infected elk tissue.
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- We dedicate the manuscript to Elizabeth Williams for
her pioneering work on CWD.
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- http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=1262585
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