Frequent coinfection with hepatitis B virus strains of distinct genotypes detected by hybridization with type-specific probes immobilized on a solid-phase support.
A genotype-specific probes assay (GSPA) was developed for distinguishing the seven genotypes (A-G) of hepatitis B virus (HBV). Nucleotide (nt) sequences corresponding to preS1 region were amplified by PCR with a primer labeled with biotin, and delivered to eight wells on which complementary sequences specific to one or other genotype had been immobilized. Thereafter, hybridization of HBV DNA sequences amplified from the test serum was detected by colorimetry. When 256 sera from HBV carriers in Bangladesh, Cameroon, Japan, South Africa, USA and Uzbekistan were subjected to GSPA, genotypes were concordant with those of ELISA with monoclonal antibodies to epitopes on preS2-region products in 242 (94.6%) of them; 8 sera (3.1%) were not genotypeable by either method. Cloning analysis confirmed the presence of two distinct HBV genotypes in the seven selected sera with coinfection. There were 7 (2.7%) sera with discordant genotyping results between GSPA and ELISA. When HBV DNA clones propagated from these sera were sequenced and analyzed phylogenetically, the genotypes determined by GSPA were verified. Coinfection with HBV strains of two distinct genotypes was identified by GSPA in 28 (10.9%) sera, while it was suggested by ELISA in only 2 (0.8%) sera. The GSPA method would be particularly useful for detecting the coinfection with distinct HBV genotypes of any clinical relevance, which seems to be more frequent than reported previously.
Kato
Hideaki
H
Department of Clinical Molecular Informative Medicine, Nagoya City University Graduate School of Medical Sciences, 467-8601, Nagoya, Japan.
Orito
Etsuro
E
Sugauchi
Fuminaka
F
Ueda
Ryuzo
R
Koshizaka
Takuya
T
Yanaka
Shoko
S
Gish
Robert G
RG
Kurbanov
Fuat
F
Ruzibakiev
Ruslan
R
Kramvis
Anna
A
Kew
Michael C
MC
Ahmad
Nooruddin
N
Khan
Mobin
M
Usuda
Sadakazu
S
Miyakawa
Yuzo
Y
Mizokami
Masashi
M
D001710:Biotin; D004797:Enzyme-Linked Immunosorbent Assay; D005838:Genotype; D006509:Hepatitis B / Q000150:complications* / Q000821:virology*; D006514:Hepatitis B Surface Antigens / Q000235:genetics; D006515:Hepatitis B virus / Q000145:classification* / Q000235:genetics*; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization / Q000379:methods*; D015345:Oligonucleotide Probes; D010802:Phylogeny; D016133:Polymerase Chain Reaction; D011498:Protein Precursors / Q000235:genetics; D017422:Sequence Analysis, DNA; D013045:Species Specificity
D006514:Hepatitis B Surface Antigens; D015345:Oligonucleotide Probes; D011498:Protein Precursors; C050806:presurface protein 1, hepatitis B surface antigen; D001710:Biotin