Occult hepatitis B virus infection: influence of S protein variants

Abstract

Background: In occult hepatitis B viral infection (OBI), the persistence of hepatitis B virus (HBV) DNA is associated with a lack of hepatitis B surface antigen (HBsAg). To assess the possible role of HBsAg immune escape variants in OBI patients, variability in the HBV S gene was evaluated for OBI patients as well as chronic HBV infection patients from the same families.

Methods: We selected 17 HBV DNA-positive/HBsAg-negative patients (OBI group) and 15 HBV DNA- and HBsAg-positive patients from OBI families (control group). The S gene was amplified and cloned, and at least 15 clones per patient were sequenced and analyzed.

Results: Although the incidence of stop codon mutations within the S region was higher in the OBI group (13.6 %) than in the control group (1.5 %, P < 0.001), this type of mutation, together with insertion and deletion mutations, was prevalent in only three OBI patients. In the major hydrophilic region (MHR), a median of 0.75 residues were altered in every 100 residues for the OBI patients, whereas 0.95 out of 100 residues were changed in the control group (P = 0.428). Furthermore, some variants that are generally considered immune escape variants, such as mutations at positions s145, s147, and s123, were only observed in less than 5 % of all the clones sequenced, in either OBI or control group.

Conclusions: Our data suggest that HBsAg variants may not play a major role in OBI pathogenesis.

Fig. 1

Distribution of OBI and control patients in 15 families included in this study. Symbols: square, male; circle, female; O, OBI patient; C, control patient; CHI, chronic HBV infection patient; ND, not detected. Each family was assigned an ID number. If a family had two OBI or control patients, a, b were used to specify them respectively

Fig. 2

Frequencies of clones with stop codon, insertion, deletion and reading frame shift mutations in the full-length S gene for OBI group (black bars) and Control group (white bars). Statistical significance was calculated by using the ANOVA test. *, P < 0.05

Fig. 3

a. Numbers of residue changes in clones from OBI group and Control group patients within the “α” determinant of the MHR. The numbers for the 260 clones (circles) in OBI group and the 259 clones (squares) in Control group are distributed according to the numbers of residue changes in the “α” determinant. Lines and numbers above them in parentheses, means of amino acid mutation/clone in the “α” determinant; numbers on the right side of circles or squares, numbers of clones. Statistical significance was calculated using a Mann–Whitney test. b. Frequencies of residue substitutions within the S protein in isolated clones from HBsAg- negative patients (OBI group, black bars, n = 17) and HBsAg- positive patients (Control group, white bars, n = 15), analyzed in intervals of 10 amino acids each. Each bar represents the percentage of mutated residues for all clones at each interval of 10 amino acids per group. *, P < 0.05