1. Production and characterization of a Brazilian candidate antigen for Hepatitis E Virus genotype 3 diagnosis.
- Author
-
de Almeida Ramos D, Miani M, Pandolfi R, Tondo L, Colli ML, Rosado Spilki F, Rovaris Gardinali N, Alves Pinto M, Kreutz LC, and Frandoloso R
- Subjects
- Amino Acid Sequence, Animals, Base Sequence, Brazil, Cloning, Molecular, Escherichia coli genetics, Escherichia coli metabolism, Hepatitis E virus classification, Hepatitis E virus genetics, Humans, Molecular Sequence Data, Polymerase Chain Reaction, RNA, Viral genetics, RNA, Viral isolation & purification, Rats, Rats, Wistar, Sequence Alignment, Sequence Analysis, RNA, Swine, Swine Diseases, Antigens, Viral immunology, Feces virology, Hepatitis E diagnosis, Hepatitis E virus immunology, Open Reading Frames genetics
- Abstract
Hepatitis E, caused by hepatitis E virus (HEV), is a viral infectious pathology of great importance in the public health. Hepatitis E outbreaks were registered in developing countries with poor or no sanitation, where drinking water was contaminated with fecal material, but also in many industrialized countries probably due to consumption of HEV-positive swine meat. In this study, we present the development and characterization of a recombinant antigen from ORF2 HEV genotype 3. Viral RNA was extracted from swine feces infected with the native virus. A total of 267 residues from the C-terminal ORF2((394-661)) coding sequence were cloned into the pET20a vector and expressed in Escherichia coli ER2566. Recombinant protein was purified by liquid chromatography and the fragment obtained a 98% homology against other human or swine HEV genotype 3 ORF2 sequences. Wistar rats were inoculated with ORF2p, developing antibodies able to recognize both the homologous antigen and the native HEV genotype 3 ORF2 present in infected stool. In parallel, HEV-negative swine were experimentally challenged with HEV genotype 3. ORF2 was detected by PCR 14 days post-inoculation in three-fourth piglets' feces and one week later by dot blot. In conclusion, this study proved the immunogenic and antigenic properties of the recombinant protein ORF2p., (© FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)
- Published
- 2016
- Full Text
- View/download PDF