Identification and characterization of a novel replicative intermediate of heron hepatitis B virus

We have identified and characterized a novel intracellular DNA replicative intermediate that is synthesized by heron hepatitis B virus (HHBV) and not by other avian hepadnaviruses. The new DNA form is synthesized in all host cells tested. The HHBV nucleic acid template, and not HHBV proteins, is responsible for the formation of the new form. The new form is comprised of a full-length minus-strand DNA and an incomplete plus-strand DNA whose 5' ends are mapped to DR2, predominantly. The 3' ends of its plus-strand are located between nucleotides 946 and 1046. Genetic analysis indicates that the sequences responsible for the formation of the new form lie between nucleotides 910 and 1364. The endogenous polymerase activity of capsids isolated from cells converted the new form into RC DNA. Intracellular capsids containing the new form are secreted inefficiently as virions, in comparison to RC- and DL DNA-containing capsids. Our analysis suggests that the new form is an incomplete RC DNA molecule that is due to a specific block or pause in the synthesis of plus-strand DNA. Our analysis also suggests that capsids become competent for efficient secretion sometime after the synthesis of 1500 nucleotides of plus-strand DNA.