Detection of hepatitis B precore mutants by the fluorescent linear polymerase chain reaction sequencing method.
To evaluate the presence of precore mutants, serum samples from 25 patients with chronic hepatitis B, HBV-DNA positive (5 HBeAg and 20 anti-HBe positive) were studied. The complete precore-core region of HBV-DNA was directly sequenced after polymerase chain reaction amplification by a fluorescent linear polymerase chain reaction sequencing method. Precore variants were detected in one HBeAg positive and in all 20 anti-HBe positive patients: in 19 cases, G to A at position 1896, with or without the substitution G to A at position 1899, in two cases C to T substitution at position 1817 which also produces a stop codon (CAA to TAA), one accompanied by the mutation G to A at position 1896. The only mutation observed in the patient who was initially HBeAg positive patient was a G to A substitution at position 1899. Consecutive serum samples from a patient with chronic hepatitis B, initially had the simultaneous presence of wild type and variant strains. Elimination of the wild-type strain was associated with reactivation of liver disease. Analysis of the sequences obtained demonstrated the heterogeneity of the hepatitis B virus genome in the precore-core region. These results indicate that the main cause of non-expression of HBeAg in chronic hepatitis B in our country is the substitution of G to A at nucleotide 1896, alone or accompanied by other variants. Fluorescent linear polymerase chain reaction is a fast and sensitive method to study heterogeneity in the precore-core region.
Rodriguez-Frias
F
F
Department of Hepatology, Hospital General Universitario Vall d'Hebron, Barcelona, Spain.
Buti
M
M
Arranz
J A
JA
Cotrina
M
M
Jardi
R
R
Esteban
R
R
Guardia
J
J
D000595:Amino Acid Sequence; D001483:Base Sequence; D004279:DNA, Viral / Q000032:analysis; D006508:Hepatitis Antibodies / Q000032:analysis; D006509:Hepatitis B / Q000235:genetics*; D006512:Hepatitis B Core Antigens / Q000235:genetics*; D006513:Hepatitis B e Antigens / Q000032:analysis / Q000276:immunology; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D015345:Oligonucleotide Probes / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*
D004279:DNA, Viral; D006508:Hepatitis Antibodies; D006512:Hepatitis B Core Antigens; D006513:Hepatitis B e Antigens; D015345:Oligonucleotide Probes