DNA of hepatitis B virus (HBV DNA) in sera from HBeAg-positive carriers is now the most important and reliable marker of infectivity, but its significance in the progression of chronic hepatitis in anti-HBe carrier status is still under discussion. In this study, viral DNA was tested by a simplified spot hybridization method in sera of 206 HBeAg-negative Italian subjects. In a group of 153 HBsAg carriers, we found that 15.6% of anti-HBe-positive and 10.5% of anti-HBe-negative samples contained viral DNA. No HBV DNA was revealed in 38 HBsAg-negative nor in 15 anti-HBs-positive subjects with different serological markers of HBV. Viral DNA in sera of HBeAg-negative patients with severe chronic liver disease was correlated with increased alaninetransferase activity and IgM anti-HBc. Thus the presence of HBV DNA in these sera not only predicts which subjects are potentially infectious but also indicates chronic progression of hepatitis. Finally, viral DNA extracted from Dane particles of nine anti-HBe-positive sera was characterized by the Southern blot technique. The hybridization pattern shows bands indicating the presence of replicative intermediates.