A 7-month-old dog was presented to the Veterinary Teaching Hospital of the Universidade Estadual de Londrina, Paraná, Brazil with a 1-week history of seizure-like activity and compulsive walking. Neurological deficits included seizures, nystagmus, absence of a menace reaction, depressed postural reactions, spastic tetraparesis, opisthotonos, and spasticity of the thoracic limbs. Cerebrospinal fluid (CSF) evaluation showed severe lymphocytic pleocytosis (554 cells/microL, with 70% lymphocytes) and a protein concentration of 17 mg/dL. The histopathologic findings in cerebrum, cerebellum, and brainstem obtained at necropsy were compatible with acute encephalomyelitis caused by canine distemper virus (CDV). Using reverse transcription-polymerase chain reaction (RT-PCR), CDV RNA was detected in both CSF and fragments of fresh brain tissue. The results indicated that CDV was the agent responsible for the clinical and laboratory presentation. Severe pleocytosis with lymphocyte predominance is an unusual finding in canine distemper and must be differentiated from granulomatous meningoencephalitis. RT-PCR on CSF is a useful, fast, and specific method to diagnose CDV infection in dogs.