Zoonotic diseases are often caused by viruses or bacteria, which adapt to new species with changes to their nucleic acid sequence. The etiologic agent of transmissible spongiform encephalopathies (TSEs), the prion, is a self-templating, abnormal isomer of a normal protein, PrPC, and does not include nucleic acids. Despite its protein-only composition, prions are transmissible and can adapt to new species. We are particularly interested in chronic wasting disease (CWD), the TSE of cervids. CWD is horizontally transmissible among cervids and has spread across much of North America and to Europe and Asia. It is apparent that excreta from CWD(+) deer is infectious, but the mechanism of horizontal transmission of CWD has not been explained. We hypothesized that deer accumulate prions in many tissues and that accumulation of prions is dictated by PrPC expression. Next, we used a battery of in vitro systems to compare the biochemical characteristics and infectivity of prions in lymphoid tissues. Finally, we hypothesized that in vitro detection of prions in saliva is hampered by the presence of an inhibitor. We demonstrated that prions accumulate in many tissues in deer and that accumulation is determined by tissue type, not PrPC expression. We confirmed that the prions in lymph nodes are infectious in vitro. Last, we confirmed the presence of an inhibitor in saliva which results in the underestimation of prion shedding in saliva. The TSE of cattle, bovine spongiform encephalopathy (BSE), crossed the species barrier and infected humans, confirming that TSEs can infect new species. However, the zoonotic potential of CWD remains unclear. For a prion from one species to infect a new host, the invading prion and the PrPC of the new host ¬must be compatible. We hypothesized that prions are most compatible with their own species' PrPC and that human PrPC could not be induced to misfold by CWD prions. Upon infection of a new host, CWD adapted, but BSE did not. Curiously, CWD prions efficiently induced the misfolding of human PrPC. We concluded that the species barrier between humans and CWD prions is not due to incompatibility of human PrPC and CWD prions. Finally, we tested a specific region of PrPC, the amino-terminal domain (NTD), for its role in the species barrier. We demonstrated that the NTD of PrPC hindered misfolding for most species, but that interactions of the NTD with the rest of molecule facilitated the misfolding of human PrPC by CWD prions. We propose that horizontal transmission among cervids is facilitated by the widespread propagation and deposition of prions in tissues. We hypothesize that prions in the periphery are infectious and that they are the source of excreted prions to which naïve cervids are exposed. We conclude that the species barrier preventing transmission of CWD to humans is not as robust as has been suggested. The species barrier is not due to incompatibility of human PrPC and CWD prions. We suggest that CWD infection of humans would be difficult to identify because CWD adapts to new species. Our work to understand horizontal transmission and the risk of CWD prions to humans contributes to an understanding of the pathology and ecology of CWD and the prevention and identification of zoonotic events involving CWD.