The current study demonstrates the oral transmission of CWD from the brain tissues of infected white-tailed deer to reindeer (Rangifer tarandus tarandus
). Antemortem testing identified PrPCWD
in two reindeer at 13.4 mpi and both animals manifested consistent clinical symptoms and succumbed to disease by 20 months. The course of disease progression and widespread distribution of PrPCWD
in nervous and lymphoreticular systems are consistent with observations from other cervids naturally or experimentally infected with CWD, including elk, white-tailed deer, mule deer, moose and red deer 
. A natural case of CWD in reindeer or caribou has not yet been identified but given the potential overlap in habitat with infected free-ranging cervid populations and the current findings of oral transmissibility, the potential for natural transmission certainly exists.
Cervids with terminal CWD typically display PrPCWD
deposition throughout multiple organ systems which progresses from early lymphatic tissue involvement, to central and peripheral nervous tissues followed by accumulation in other tissues including the endocrine system and heart 
. In our study, RAMALT sampling prior to 13.4 mpi was not conducted precluding an accurate determination of when lymphoid involvement occurred, although PrPCWD
has been detected in peripheral lymphoid tissue as early as 42 d after oral inoculation of mule deer fawns 
. Our observations of PrPCWD
-specific staining in sections of pancreas, pituitary, adrenal medulla, lung, kidney, bladder and salivary gland corroborate findings in other CWD-infected cervids 
, with PrPCWD
primarily localized to lymphoid or neural components of these tissues. The presence of PrPCWD
in reindeer excreta such as saliva, urine and feces is anticipated based on studies in deer 
but was not investigated here. Skeletal muscle from CWD-infected mule deer has been shown to contain infectivity 
was recently reported in muscle-associated nerve fascicles of white-tailed deer 
. We were unable to detect PrPCWD
in myocytes or associated nerves by IHC or ELISA consistent with the negative findings of others 
. Regardless, the widespread distribution of PrPCWD
in infected reindeer tissues merits consideration while evaluating the risks emanating from processing or consuming potentially infected animals.
None of the reindeer inoculated with CWDELK
developed clinical disease or accumulated PrPCWD
in any of the tissues tested. The apparent resistance of reindeer to infection by the CWDELK
inoculum may relate to subtle differences in the CWD strains present within the pooled elk and white-tailed deer inocula, however, we think this is unlikely. Differences between the two inocula were not evident through glycosylation profile analysis of PrPres
by western blot (data not shown). Further characterization of these inocula in transgenic mice is ongoing. We also considered that this may have been due to a low prion titre in the elk inoculum, however, the same CWD inoculum pool has been used in other studies and was highly infectious for red deer 
and elk 
. Alternatively, resistance could be due to the PRNP
genotype, in which heterozygous codons occurred at positions 2 (V/M), 129 (G/S), 138 (S/N), and 169 (V/M) in the mature form of PrPC
. An important role for PRNP
polymorphisms in determining disease susceptibility is also suggested by the finding that one of the three reindeer has not yet succumbed to infection with the CWDWTD
inoculum by 26 mpi and showed no detectable PrPCWD
in lymphoid tissue at 25 mpi. This possibly resistant reindeer differs from the susceptible reindeer in being heterozygous at a single codon 138 (S/N).
The role of codons 2, 129 and 169 in CWD resistance is unknown. Interestingly, 138 N is common to the 4 reindeer that resisted elk or white-tailed deer CWD infection and may offer some protection. Consistent with this possibility, fallow deer were reported to express a PRNP
gene encoding 138 N and 226 E and resisted infection when housed together with CWD-infected mule deer for 7 years 
. Residue 138 N was implicated as the residue possibly protective against CWD infection, since red deer that express PrPC
with 226 EE are highly susceptible to elk CWD infection 
. Fallow deer were shown to be susceptible to CWD inoculation by the intracerebral route with long incubation periods of 4–5 years 
. Nevertheless, it will be important to determine whether fallow deer resist CWD after an oral challenge.
Codon 138 is in the β1−α1 loop region 
. Although serine and asparagine are similar small, polar residues, serine/asparagine differences at position 173 (elk numbering) of the β2−α2 loop may impact the susceptibility of a species to CWD infection 
. Additionally, crystal structures of the β2−α2 loop peptides have shown that 173 (S/N) and 177 (N/T) sequence differences modify how the side chains pack at the β-sheet interface, and suggest that this incompatibility may underlie transmission barriers 
. Whether serine or asparagine at position 138 has a similar effect on β-sheet packing is not yet known.
Would the reindeer that were heterozygous at 2 (V/M), 129 (G/S), 138 (S/N), and 169 (V/M) completely resist CWD infection or require a long incubation period to develop PrPCWD or clinical signs of disease? No trace of PrPCWD in lymphoid tissue or any other tissues at the end of the experiment (60 mpi) suggests at minimum a long delay in developing CWD infection, and at best complete protection from CWD infection.
In North America, the term reindeer describes a domesticated Rangifer tarandus
subspecies (Rangifer tarandus tarandus
) introduced from Eurasia approximately a century ago, while the term caribou refers to several native wild Rangifer tarandus
subspecies. Major caribou subspecies include woodland (Rangifer tarandus caribou
) and barren-ground (Rangifer tarandus groenlandicus
, Rangifer tarandus granti
) caribou 
. Barren-ground caribou generally exist in higher northern latitudes, in some regions commingling with semi-domesticated reindeer herds, while the range of woodland caribou extends further south into the provinces of Alberta and Saskatchewan which currently harbour CWD-infected cervids. A study of 95 caribou in northern Quebec failed to identify CWD 
, although CWD has not yet been detected in Canadian provinces east of Saskatchewan.
Interbreeding between reindeer and caribou has occurred in captivity and the wild although general genetic introgression appears to be minimal 
. Two of the reindeer protected from CWD were heterozygous at 4 codons that were in common with the reported genotypes of Rangifer tarandus granti
and differ from Rangifer tarandus tarandus
(unpublished, Genbank accession #AY639093.01). It is possible that the two resistant reindeer were derived from interbreeding two Rangifer tarandus
subspecies, potentially captive and free-ranging. It will be crucial to determine the prevalence of the polymorphisms in both free-ranging and captive reindeer populations, particularly considering the possible protective PRNP
Large scale analysis of PRNP
alleles in reindeer has not been reported. Largely consistent with our findings in captive reindeer, Happ et al
recently sequenced PRNP
from three free-ranging caribou herds in Alaska and identified polymorphisms at codons 2 (V/M), 129 (G/S), 138 (S/N) and 169 (V/M). Moreover, the most frequent caribou allele across the 3 herds examined was VGSV, which is common to the 2 reindeer we found to be susceptible to CWDWTD
. The close interrelatedness of Rangifer
species implies disease susceptibility in one may predict susceptibility in the others, although a single amino acid difference in a key position could confer resistance.
The importance of Rangifer
species to the culture of aboriginal peoples cannot be underestimated with many components of hunted animals being consumed as food. Although relatively limited in comparison to elk and deer industries in North America, reindeer and caribou farming does occur, producing consumables such as meat, hides and antler velvet. The human health risks of consuming meat or other products derived from CWD-infected animals remain uncertain, although epidemiological evidence indicates transmission has not yet occurred 
and transgenic mouse studies suggest the risk is remote in humans expressing common PRNP
. The finding that CWD can be transmitted to squirrel monkeys by intracranial inoculation 
raises concern for human transmissibility, however a study in macaques failed to demonstrate transmission after 70 months 
. Since prion strains may undergo changes in transmission characteristics following passage through different species and strain selection pressures can be exerted by host genetic factors during passage within a species 
, caution is warranted when predicting the risks of CWD transmission from reindeer to other species.
This is the first evidence of CWD transmission to the sub-species Rangifer tarandus tarandus, implicating the potential for transmission to others in this genus. Current diagnostic tests, including antemortem RAMALT testing, appear capable of detecting CWD in Rangifer species and increased surveillance would be required to determine if natural transmission has indeed occurred. Additional studies are ongoing to chart the distribution of infectivity during the course of disease and determine the influence of PRNP polymorphisms in disease susceptibility.