AbstractBat virus host shifts can result in the spread of diseases with significant effects. The rabies virus (RABV) is able to infect almost all mammals and is therefore a useful model for the study of host shift mechanisms. Carnivore RABVs originated from two historical host shifts from bat viruses. To reveal the genetic pathways by which bat RABVs changed their host tropism from bats to carnivores, we investigated the second permanent bat-to-carnivore shift resulting in two carnivore variants, known as raccoon RABV (RRV) and south-central skunk RABV (SCSKV). We found that their glycoprotein (G) genes are the result of recombination between an American bat virus and a carnivore virus. This recombination allowed the bat RABV to acquire the head of the G-protein ectodomain of the carnivore virus. This region is involved in receptor recognition and binding, response to changes in the pH microenvironment, trimerization of G proteins, and cell-to-cell transmission during the viral infection. Therefore, this recombination event may have significantly improved the variant’s adaptability to carnivores, altering its host tropism and thus leading to large-scale epidemics in striped skunk and raccoon.