Sequence conservation and divergence of hepatitis delta virus RNA

The complete RNA sequence of the hepatitis delta virus (HDV) obtained from the Nauru Island in the Pacific was determined by cDNA cloning and amplification by polymerase chain reaction (PCR). The sequence showed 14-17% divergence from the two known HDV RNA sequences. There are three highly conserved domains: the region around the autocatalytic cleavage site of the genomic RNA (nucleotides 659 to 772), the region around the autocatalytic cleavage site of the antigenomic-sense RNA (nucleotides 847 to 966), and the region around the middle one-third domain of the open reading frame (ORF) encoding the hepatitis delta antigen on the antigenomic RNA (nucleotides 1267 to 1347). The two autocatalytic activities are required for the cleavage and ligation of HDV RNA during RNA replication. The third conserved domain codes for the RNA-binding domain of HDAg, which specifically interacts with HDV RNA. Three nucleotide changes within the genomic catalytic sequence are present but did not alter the catalytic cleavage activity of the HDV RNA. Microheterogeneity of the RNA sequences was also detected. One of these occurred within the coding region of the delta antigen, creating an amber termination codon in some of the RNA species. Thus, this HDV strain contains two different RNA species, one of which encodes a delta antigen of 214 amino acids and the other 195 amino acids. These two protein species were detected by immunoblotting of the patient's plasma. In contrast to other HDV strains, only three ORFs capable of encoding more than 100 amino acids each are present in this HDV RNA. We recommend that oligonucleotides complementary to the highly conserved sequences should be used as primers for PCR in clinical detection assays of hepatitis delta virus infection.