Rapid detection and further characterization of infection with hepatitis B virus variants containing a stop codon in the distal pre-C region.
Recently, hepatitis B virus (HBV) replication in the absence of HBe antigenaemia has been attributed to HBV variants with a TAG stop codon in the distal pre-C region associated with one or two point mutations. We describe here a rapid detection method for the diagnosis of such HBeAg-negative HBV variants using selective oligonucleotide hybridization. The entire pre-C region was amplified by the polymerase chain reaction and hybridized under stringent conditions with non-mutated (M0), one (M1) and two (M2) point-mutated oligonucleotide probes. Of the 15 HBeAg-positive (group I) and 20 HBeAg-negative (group II) serum samples studied, 14 samples in group I and one sample in group II hybridized with M0 only and 18 samples in group II hybridized with M1 or M2, or both. The remaining two samples (from groups I and II, respectively) failed to hybridize with any of the three probes. DNA sequencing confirmed mixed distal pre-C sequences in samples hybridizing with more than one probe and also revealed novel mutations in the distal pre-C region of the two samples which failed to hybridize with any of the probes. The latter sample had a +2 frameshift and hence represented a new type of HBeAg-negative HBV variant. This method may therefore prove useful in the diagnosis of infections by HBeAg-negative HBV variants resulting from common mutations in the pre-C region, as well as for the identification of less common variants with novel mutations in the same region.
Unité de Recherche sur les Hepatites, le SIDA et les Rétrovirus Humains INSERM 271, Lyon, France.
J
J
Li
S
S
Tong
L
L
Vitvitski
F
F
Zoulim
C
C
Trépo
D001483:Base Sequence; D003062:Codon / Q000235:genetics*; D004279:DNA, Viral / Q000235:genetics* / Q000302:isolation & purification; D014644:Genetic Variation; D006509:Hepatitis B / Q000175:diagnosis*; D006513:Hepatitis B e Antigens / Q000032:analysis / Q000235:genetics*; D006515:Hepatitis B virus / Q000235:genetics* / Q000302:isolation & purification; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D009693:Nucleic Acid Hybridization; D015345:Oligonucleotide Probes; D016133:Polymerase Chain Reaction
D003062:Codon; D004279:DNA, Viral; D006513:Hepatitis B e Antigens; D015345:Oligonucleotide Probes