This research work was conducted to find out nucleotide variations in the pre-S2 region of the two clinically diagnosed Surface Antigen of Hepatitis B Virus (HBsAg). Pre-S2 region nucleotide sequence of the sample MR02 was found completely homologous with the Pre-S2 region of already reported subtype “adr4”. This sample showed 1.2 to 20% variation with the other reported sequences. Nucleotide sequence of the Pre-S2 region of the second sample MR03 does not show 100% homology with any of the reported sequences. It differs 2.4 to `18.8 % from the other reported sequences. Further it was established that MR02 and MR03 nucleotide sequences differ from one another at four positions, all substitutions. The first difference is observed at 87th base where there is “C” residue in MR02 DNA sequence and an “A” in MR03 DNA sequence. The 2nd, 3rd and 4th difference is at base number 116, 137 and 146 respectively. In all the cases “C” residue in MR02 sequence is replaced by “T” residue in MR03 nucleotide sequence. The first change at nucleotide level does not cause any change at amino acid level, while the 2nd, 3rd and 4th substitutions at nucleotide level results an amino acid change. The phylogenetic studies established that the Pakistanian HBV Isolates closely resemble the “adrq” strain belonging to “adr” subtype. The study showed that the pre-S2 region is hypervariable and two reported sequences show 100% homology and thus cause change in the structure of Hepatitis B Surface Antigen (HBsAg). This variability reduces the efficacy of vaccines prepared to a particular clone of HBV isolates.
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