Canine distemper virus (CDV) is responsible for high morbidity and mortality in dogs worldwide. Epidemiological study of canine distemper can help to control and treat the disease in any area. This study aimed to investigate the prevalence of CDV in dogs referred to the Veterinary Hospital from September 23, 2018 to September 22, 2019. Dogs with at least two clinical signs of canine distemper underwent blood tests, rapid test kit from the eye and cerebrospinal fluid (CSF), and RT-PCR from whole blood and/or CSF samples. Out of 1212 referred dogs, 112 dogs were suspected to have canine distemper of which 90 underwent RT-PCR and rapid test kits. The disease prevalence was 4.04% (49/1212) and 7.44% (49/659) according to the total number of referring dogs and number of referring sick dogs, respectively. The distemper fatality rate was 69.57% (32/46). Seventy percent of distemper positive cases were under 12 months old and 52.08% were under 6 months old. Female dogs were more susceptible than males; however, the fatality rate of males was more than females. Of distemper positive dogs, 91.84% were unvaccinated. The highest prevalence (71.43%) of dogs diagnosed with CDV occurred during the cold seasons. It is concluded that canine distemper is endemic in the geographical area of Mashhad and its prevalence rate in dogs referred to the Veterinary Hospital of Ferdowsi University of Mashhad is 4.04% and its fatality rate is 69.57%. This indicates that a significant number of dogs may die if they develop distemper despite treatment.