The first human virus in the genus Cardiovirus was described in 2007 and named Saffold virus (SAFV). Cardioviruses can cause severe infections of the myocardium and central nervous system in animals, but SAFV has not yet been convincingly associated with disease in humans. To study a possible association between SAFV and infections in the human central nervous system, we designed a real-time PCR for SAFV and tested cerebrospinal fluid (CSF) samples from children <4 years of age. SAFV was detected in 2 children: in the CSF and a fecal sample from 1 child with monosymptomatic ataxia caused by cerebellitis; and in the CSF, blood, and myocardium of another child who died suddenly with no history of illness. Virus from each child was sequenced and shown to be SAFV type 2. These findings demonstrate that SAFV can cause serious invasive infection in children.