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.
Li
J
J
Unité de Recherche sur les Hepatites, le SIDA et les Rétrovirus Humains INSERM 271, Lyon, France.
Tong
S
S
Vitvitski
L
L
Zoulim
F
F
Trépo
C
C
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