PreS1 Mutations Alter the Large HBsAg Antigenicity of a Hepatitis B Virus Strain Isolated in Bangladesh

Abstract

Mutations in the hepatitis B virus (HBV) genome can potentially lead to vaccination failure, diagnostic escape, and disease progression. However, there are no reports on viral gene expression and large hepatitis B surface antigen (HBsAg) antigenicity alterations due to mutations in HBV isolated from a Bangladeshi population. Here, we sequenced the full genome of the HBV isolated from a clinically infected patient in Bangladesh. The open reading frames (ORFs) (P, S, C, and X) of the isolated HBV strain were successfully amplified and cloned into a mammalian expression vector. The HBV isolate was identified as genotype C (sub-genotype C2), serotype adr, and evolutionarily related to strains isolated in Indonesia, Malaysia, and China. Clinically significant mutations, such as preS1 C2964A, reverse transcriptase domain I91L, and small HBsAg N3S, were identified. The viral P, S, C, and X genes were expressed in HEK-293T and HepG2 cells by transient transfection with a native subcellular distribution pattern analyzed by immunofluorescence assay. Western blotting of large HBsAg using preS1 antibody showed no staining, and preS1 ELISA showed a significant reduction in reactivity due to amino acid mutations. This mutated preS1 sequence has been identified in several Asian countries. To our knowledge, this is the first report investigating changes in large HBsAg antigenicity due to preS1 mutations.

Impact of the Interaction of Hepatitis B Virus with Mitochondria and Associated Proteins

“…Its genome encodes four overlapping open reading frames (ORFs) known as polymerase (pol; P), surface or envelopes (S), core (C), and X protein (X) (Figure 1) [11][12][13][14]. HBV enters hepatocytes through sodium taurocholate cotransporting polypeptide (NTCP) as a receptor to bind with the preS1 region in the envelope protein, and then the uncoated nucleocapsid is transported to the nucleus, where the relaxed circular DNA (rcDNA) genome is converted to covalently closed circular DNA (cccDNA) [15][16][17]. The cccDNA is competent for transcription of 3.5 kb pregenomic RNA (pgRNA), and for transcription of several subgenomic RNAs such as 2.4 kb preS-S mRNA, 2.1 kb S mRNA, and 0.7 kb X mRNA.…”

Complete Genome Sequence of a Precore-Defective Hepatitis B Virus Genotype D2 Strain Isolated in Bangladesh

“…H epatitis B virus (HBV), a causative agent for acute/chronic hepatitis and hepatocellular carcinoma (HCC), has a circular partially double-stranded DNA genome, is about 3.2 kb long, and belongs to the Hepadnaviridae family and Orthohepadnavirus genus (1,2). Nucleotide sequence variation in the genome has been used to classify HBV into different genotypes (A to J) (3,4). However, the prevalences of HBV genotypes and subtypes vary depending on the geographical region, and genomic variations have been associated with HBV pathogenesis (2,5).…”

“…HBV was detected in a patient serum sample (3). Viral DNA from serum samples was extracted using a QIAamp DNA minikit according to the manufacturer's protocol.…”

“…Viral DNA from serum samples was extracted using a QIAamp DNA minikit according to the manufacturer's protocol. Amplification of the HBV complete overlapping ORFs P, pre-C, and X and the partial S ORF was carried out using specific primers (Table 1), as previously described (3). The PCR products were purified using a MonoFas DNA purification kit (GL Sciences, Inc., Tokyo, Japan) and were used as the templates for sequencing.…”

“…The PCR products were purified using a MonoFas DNA purification kit (GL Sciences, Inc., Tokyo, Japan) and were used as the templates for sequencing. The sequencing reactions were performed according to our previously reported protocol using the BigDye (3). Raw sequence data were edited, annotated, and analyzed using CLC Sequence Viewer.…”