ABSTRACT Prion diseases are a group of infectious neurodegenerative disorders affecting humans and other mammals, which are transmitted by an unconventional proteinaceous infectious agent termed prion. CWD is so far the only prion disease of wild animals; it is highly contagious and the exact origin, prevalence, and mechanisms of transmission remain incompletely understood. The progressive increase of CWD prevalence in the USA has likely resulted in a substantial contamination of the environment with infectious prions and may lead to new prion diseases in animals that share the habitat of cervids. The disease has been rapidly expanding geographically and now affects 27 states in USA, three Canadian provinces, South Korea and Northern Europe, including Norway, Sweden and Finland. The risk of transmission of CWD to other animal species or to humans is unknown and surveillance methods to detect the infection in non-cervid species are limited. This Program Project has received continuous funding for the past 11 years, which has enabled us to implement a large and complementary set of unique techniques, model systems, reagents, personnel and expertise crucial to understand the CWD problem. In the last competitive review, we received a perfect score of 10 and reviewers identified many strengths and “not any significant weakness”. The progress during this cycle has been stellar, with many important discoveries and 118 high profile publications. The main goal of this iteration of the Program Project will be to study the molecular and cellular mechanisms responsible for prion replication and prion strain diversity in CWD. We will study the generation, mutation and evolution of CWD prion strains, the inter-species transmission potential of CWD (with particular focus on its zoonotic potential) and the role of the environment on CWD transmission. This Program Project aims to address some of the most important questions in CWD research, including the possibly origin of CWD, the mechanism and routes of transmission of the disease among animals, the natural strain diversity of CWD, the atomic resolution structure of infectious prions, the evaluation of the zoonotic potential of CWD, the factors controlling the cervid/human species barrier, the role of the environment on prion transmission and the development of efficient procedures to detect infectious CWD prions. To achieve these goals, this Program Project assembles a group of highly accomplished scientists in the prion field, particularly experts on CWD transmission, who have access to the relevant models, techniques, samples, resources and expertise to provide a complementary and comprehensive investigation of the problem.