Molecular diversity of the E2/NS1 gene and 5'UT region of the Hepatitis C virus genome

Abstract

Individuals infected with the hepatitis C virus (HCV), while undergoing alpha-interferon (α-IFN) treatment, either respond to therapy or relapse into a chronic state. The low overall response rate to α-IFN has been suggested to result from genetic heterogeneity in the HCV genome (Major and Feinstone 1997). This non-responsiveness makes use of pre-treatment predictive indicators essential, including the correct genotyping of HCV strains.

To determine the role of genetic diversity in the HCV's resistance to α-IFN treatment, reverse transcription polymerase chain reaction was performed on sixteen New Zealand isolates, amplifying the envelope 2/non-structural (E2/NS1) region and fifty patients from the relatively conserved 5’untranslated (5'UT) region. Only two of the sixteen E2/NS1 patients were sequenced and aligned with eight overseas sequences, revealing local sequences to be distinct from the overseas strains. Amino acid comparison supported these findings identifying a hypervariable region with nonsynonymous nucleotide substitutions. The remaining fourteen patients yielded inconsistent PCR products.

Twenty-one different local 5'UT region sequences were identified from the fifty chronically infected individuals analysed. Four were subtype 1a, three were 1a-variant, seven were 1b, ten were 1b-variant, two were 2a, eight were 2b, twelve were 3a and four were 4a. Sequence polymorphism identified may alter the secondary structure of the 5'UT region. Sequence comparison revealed New Zealand strains to be closely related to Japanese strains. The extent of sequence variation observed in the New Zealand isolates may indicate the length of time HCV has existed locally. Increased genetic diversity over time in the dominant New Zealand strains, 1bv and 3a, may also exacerbate the observed unresponsiveness to α-IFN and therefore affect clinical management.

The differences in the restriction sites that are used to classify HCV subtypes do not necessarily represent sequence homology between strains, illustrated by two individuals classified by conventional restriction fragment length polymorphism, who were re-classified from our sequencing data, therefore indicating sequencing as a more precise classification method.