The aim of this study was to determine the prevalence of hepatitis viruses and analyze the epidemiological features in acute hepatitis in Spain. Consecutive serum samples from 341 patients with acute hepatitis were tested for IgM anti-hepatitis A virus, IgM anti-hepatitis B core antigen and HBsAg. The antibody to hepatitis C virus was determined using a second-generation enzyme immunoassay. In selected cases, serum was tested for antibodies for hepatitis E virus by enzyme immunoassay and hepatitis C virus-RNA by polymerase chain reaction. One hundred and fourteen (33.2%) cases of acute hepatitis were due to hepatitis A virus, 68 (20%) to hepatitis B virus, 21 (6%) to hepatitis D virus and 74 (21.8%) to hepatitis C virus infection. Of the 74 hepatitis C virus infections, 48 (65%) were anti-HCV positive at the time of diagnosis, while 26 (35%) seroconverted to anti-HCV later during follow-up. No case of hepatitis E virus infection was detected. The remaining 64 cases were negative to all acute serological markers and were classified as non-A, non-B, non-C, non-E hepatitis. Hepatitis C virus was implicated in 65/119 (54.6%) of the cases among intravenous drug users but in only in 9/199 (4.5%) of the sporadic cases (p < 0.01). Progression to chronic hepatitis was observed more frequently in anti-hepatitis C virus-positive than in antibody-negative-cases (42/70; 60% vs. 3/52 or 5.5%) (p < 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)