Persistent and transient antibody responses to hepatitis E virus detected by western immunoblot using open reading frame 2 and 3 and glutathione S-transferase fusion proteins.
Recombinant proteins containing amino acid sequences from open reading frame (ORF) 2 and ORF3 of a Chinese strain of hepatitis E virus (HEV) were constructed as fusions with glutathione S-transferase (GST). Stable fusion proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the proteins were transferred to nitrocellulose membranes, and the immobilized proteins were probed with sera from hepatitis E patients from various regions or from rhesus monkeys (Macaca mulatta) experimentally infected with the Chinese strain of HEV. Immunoglobulin G-class antibodies were detected with chemiluminescence and anti-human immunoglobulin G conjugated to horseradish peroxidase. Anti-ORF3 antibodies were detected in most patients and monkeys within 17 days of exposure, but this humoral response declined with time and was usually undetectable by approximately 100 days. Anti-ORF2.1 antibody was usually detected as early as anti-ORF3 but persisted in all animals and many patients, whereas reactivity to the larger GST-ORF2.2 fusion protein was more transient, even though all sequences present in GST-ORF2.1 are present in GST-ORF2.2. Rechallenge of these monkeys with HEV suggested that immunity to reinfection was incomplete, as levels of anti-ORF2.1 (but not anti-ORF2.2) were boosted after each rechallenge. The results demonstrate that the carboxy-terminal region of HEV ORF2 contains epitopes which are recognized by convalescent-phase antibody and are likely to be associated with limited immunity to infection, but these epitopes may be masked when larger portions of ORF2 are expressed as recombinant proteins.
Li
F
F
Macfarlane Burnet Centre for Medical Research, Melbourne, Australia.
Zhuang
H
H
Kolivas
S
S
Locarnini
S A
SA
Anderson
D A
DA
D000818:Animals; D000956:Antigens, Viral / Q000235:genetics / Q000276:immunology*; D001483:Base Sequence; D015153:Blotting, Western; D000939:Epitopes / Q000276:immunology; D005982:Glutathione Transferase / Q000235:genetics; D006508:Hepatitis Antibodies / Q000096:biosynthesis* / Q000097:blood / Q000276:immunology; D016751:Hepatitis E / Q000097:blood / Q000276:immunology; D016752:Hepatitis E virus / Q000235:genetics / Q000276:immunology*; D006801:Humans; D008253:Macaca mulatta; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D012367:RNA, Viral / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000276:immunology*; D014764:Viral Proteins / Q000235:genetics / Q000276:immunology*
D000956:Antigens, Viral; D000939:Epitopes; D006508:Hepatitis Antibodies; D012367:RNA, Viral; D011993:Recombinant Fusion Proteins; D014764:Viral Proteins; D005982:Glutathione Transferase