Artificial mosaic protein containing antigenic epitopes of hepatitis E virus.

We recently identified a novel virus, designated avian hepatitis E virus (avian HEV), from chickens with hepatitis–splenomegaly (HS) syndrome in the USA. We showed that avian HEV is genetically related to swine and human HEVs. Here we report the antigenic cross-reactivity of the putative open reading frame 2 (ORF2) capsid protein of avian HEV with those of swine and human HEVs and the Australian chicken big liver and spleen disease virus (BLSV). The region encoding the C-terminal 268 amino acid residues of avian HEV ORF2 was cloned into expression vector pRSET-C. The truncated ORF2 protein was expressed in E. coli as a fusion protein and purified by affinity chromatography. Western blot analysis revealed that the avian HEV ORF2 protein reacted with antisera against the Sar-55 strain of human HEV and with convalescent antisera against swine HEV and the US2 strain of human HEV, as well as with antiserum against BLSV. Convalescent sera from specific-pathogen-free chickens experimentally infected with avian HEV also reacted with the recombinant capsid proteins of swine HEV and Sar-55 human HEV. Antisera against the US2 human HEV also reacted with recombinant ORF2 proteins of both swine HEV and Sar-55 human HEV. The antigenic cross-reactivity of the avian HEV putative capsid protein with those of swine and human HEVs was further confirmed, for the most part, by ELISA assays. The data indicate that avian HEV shares certain antigenic epitopes in its putative capsid protein with swine and human HEVs, as well as with BLSV. The results have implications for HEV diagnosis and taxonomy.

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Antigenic Domains of the Open Reading Frame 2-Encoded Protein of Hepatitis E Virus

Journal of Clinical Microbiology ◽

10.1128/jcm.37.9.2863-2871.1999 ◽

1999 ◽

Vol 37 (9) ◽

pp. 2863-2871 ◽

Cited By ~ 38

Author(s):

Yury E. Khudyakov ◽

Elena N. Lopareva ◽

Danny L. Jue ◽

Tamara K. Crews ◽

S. P. Thyagarajan ◽

...

Keyword(s):

Synthetic Peptides ◽

Hepatitis E Virus ◽

Hepatitis E ◽

Open Reading Frame ◽

Reading Frame ◽

Orf2 Protein ◽

Positive Serum ◽

Antigenic Domains ◽

Open Reading Frame 2 ◽

Antigenic Epitopes

The antigenic composition of the hepatitis E virus (HEV) protein encoded by open reading frame 2 (ORF2) was determined by using synthetic peptides. Three sets of overlapping 18-, 25-, and 30-mer peptides, with each set spanning the entire ORF2 protein of the HEV Burma strain, were synthesized. All synthetic peptides were tested by enzyme immunoassay against a panel of 32 anti-HEV-positive serum specimens obtained from acutely HEV-infected persons. Six antigenic domains within the ORF2 protein were identified. Domains 1 and 6 located at the N and C termini of the ORF2 protein, respectively, contain strong immunoglobulin G (IgG) and IgM antigenic epitopes that can be efficiently modeled with peptides of different sizes. In contrast, antigenic epitopes identified within the two central domains (3 and 4) were modeled more efficiently with 30-mer peptides than with either 18- or 25-mers. Domain 2 located at amino acids (aa) 143 to 222 was modeled best with 25-mer peptides. A few 30-mer synthetic peptides derived from domain 5 identified at aa 490 to 579 demonstrated strong IgM antigenic reactivity. Several 30-mer synthetic peptides derived from domains 1, 4, and 6 immunoreacted with IgG or IgM with more than 70% of anti-HEV-positive serum specimens. Thus, the results of this study demonstrate the existence of six diagnostically relevant antigenic domains within the HEV ORF2 protein.

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