An outbreak of acute hepatitis of unknown aetiology in children was first reported in Scotland in April 2022. We carried out a detailed investigation of nine cases and 58 control subjects. Using next-generation sequencing and real-time polymerase chain reaction, our team compared samples and were able to confirm the presence of AAV2 in the plasma and liver of all nine cases. There was no AAV2 in any of the subjects in the control groups, which were made up of: age-matched healthy controls; children with adenovirus but normal liver function; and children admitted to hospital with known causes of hepatitis. AAV2 is not known to normally cause disease and often accompanies infection with adenoviruse, but we have now shown presence of the AAV2 virus associated with unexplained hepatitis in children. Larger studies are urgently needed to investigate the unanswered questions about the role of AAV2 in paediatric hepatitis cases.