Expression and Immunoreactivities of Hepatitis E Virus Genotype 3 Open Reading Frame-2 (ORF-2) Recombinant Proteins Expressed in Insect Cells
Hepatitis E virus (HEV) is a fecal-orally transmitted virus that is endemic in many geographical areas with poor sanitary conditions and inadequate water supplies. In Europe, a low-endemic area, an increased number of autochthonous sporadic human cases of patients infected with HEV strains of genotype 3, have been reported lately. The relatively high prevalence of HEV genotype 3 infections in European pigs has raised concerns about a potential zoonotic transmission to humans. Determination of HEV seroprevalence in pigs would help to clarify its incidence and possible zoonotic implications. To this purpose, we have expressed and partially characterized swine genotype 3 HEV open reading frame-2 proteins upon infection of Sf21 insect cells with recombinant baculoviruses. The use of the expressed proteins as diagnostic antigens for the detection of antibodies to HEV has been further assayed with human and swine sera.
KeywordsZoonosis Emerging virus Immunogenicity ELISA
The study was supported in part by grants (AGL2004-06071 and CSD2006-0007) from the Spanish Ministerio de Ciencia e Innovación (MICINN) to J.C.S. N.J.O. has been supported by a scholarship from Instituto Nacional de Investigación y Tecnología Agraraia y Alimentaria (INIA) and by a COST-929 short-term mission. J.A. was supported by a scholarship from the MICIIN. E.E.R. was supported by a contract ascribed to grant CSD2006-0007 from MICIIN. We are indebted to M.T. Pérez-Gracia (U. Cardenal Herrera-CEU, Valencia, Spain), M. Buti (Hospital Vall d′Hebron. Barcelona, Spain), and M. Martín (CReSA, UAB, Barcelona, Spain) for kindly providing us with swine fecal extract, human and swine sera, respectively.
- Arankalle, V. A., Lole, K. S., Deshmukh, T. M., Chobe, L. P., & Gandhe, S. S. (2007). Evaluation of human (genotype 1) and swine (genotype 4)-ORF2-based ELISAs for anti-HEV IgM and IgG detection in an endemic country and search for type 4 human HEV infections. Journal of Viral Hepatitis, 14(6), 435–445.PubMedCrossRefGoogle Scholar
- Banks, M., Heath, G. S., Grieson, S. S., King, D. P., Gresahm, A., Gironés, R., et al. (2004). Evidence for the presence of hepatitis E virus in pigs in the United Kingdom. Veterinary Records, 154(8), 223–227.Google Scholar
- Borgen, K, Herremans, T., Duizer, E., Vennema, H., Rutjes, S., Bosman, A., et al. (2008). Non-travel related hepatitis E virus genotype 3 infections in the Netherlands: A case series 2004–2006. BMC Infectious Diseases. doi: 10.1186/1471-2334-8-61.
- Bouwknegt, M., Engel, B., Herremans, M. M. P. T., Widdowson, M. A., Worm, H. C., Koopmans, M. P. G., et al. (2007a). Bayesian estimation of Hepatitis E virus seroprevalence for populations with different exposure levels to swine in the Netherlands. Epidemiology and Infection, 136(4), 567–576.PubMedGoogle Scholar
- Bouwknegt, M., Frankena, K., Rutjes, S. A., Wellenberg, G. J., de Roda Husman, A. M., van der Poel, W. H., et al. (2008). Estimation of hepatitis E virus transmission among pigs due to contact exposure. Veterinary Research. doi: 10.1051/vetres:2008017.
- Casas, M., Pujols, J., Rosell, R., de Deus, N, Peralta, B., Pina, S., et al. (2008). Retrospective serological study of hepatitis E infection in pigs from 1985 to 1997 in Spain. Veterinary Microbiology. doi: 10.1016/j.vetmic.2008.09.075.
- Dalton, H. R., Stableforth, W., Thurairajah, P., Hazeldine, S., Remnarace, R., Usama, W., et al. (2008). Autochthonous hepatitis E in Southwest England: Natural history, complications and seasonal variation, and hepatitis E virus IgG seroprevalence in blood donors, the elderly and patients with chronic liver disease. European Journal of Gastroenterology and Hepatology, 20(8), 784–790.PubMedCrossRefGoogle Scholar
- Fernández-Barredo, S., Galiana, C., García, A., Vega, S., Gómez, M. T., & Pérez-Gracia, M. T. (2006). Detection of hepatitis E virus shedding in feces of pigs at different stages of production using reverse transcription polymerase chain reaction. The Journal of Veterinary Diagnostic Investigation, 18(5), 462–465.Google Scholar
- Li, F., Torresi, J., Locarnini, S. A., Zhuang, H., Zhu, W., Guo, X., et al. (1997a). Amino-terminal epitopes are exposed when full-length open reading frame 2 of hepatitis E virus is expressed in Escherichia coli, but carboxy-terminal epitopes are masked. Journal Medical Virology, 52(3), 289–300.CrossRefGoogle Scholar
- Meng, X. J., Wiseman, B., Elvinger, F., Guenette, D. K., Toth, T. E., Engle, R. E., et al. (2002). Prevalence of antibodies to hepatitis E virus in veterinarians working with swine and in normal blood donors in the United States and other countries. Journal of Clinical Microbiology, 40(1), 117–122.PubMedCrossRefGoogle Scholar
- Mizuo, H., Yazaki, Y., Sugarawa, K., Tsuda, F., Takahasi, M., Nishizawa, T., et al. (2005). Possible risk factors for the transmission of hepatitis E virus and for the severe form of hepatitis E acquired locally in Hokkaido, Japan. Journal of Medical Virology, 76(3), 341–349.PubMedCrossRefGoogle Scholar
- Pintó, R., & Saiz, J. C. (2007). Human viruses in water. In A. Bosch (Ed.), Perspectives in medical virology (Vol. 17, pp. 39–67). Amsterdam, The Netherlands: Elsevier.Google Scholar
- Robinson, R. A., Burgess, W. H., Emerson, S. U., Leibowitz, R. S., Sosnovtseva, S. A., Tsarev, S., et al. (1998). Structural characterization of recombinant hepatitis E virus ORF2 proteins in baculovirus-infected insect cells. Protein Expression and Purification, 12(1), 75–84.PubMedCrossRefGoogle Scholar
- Tsarev, S. A., Tsareva, T. S., Emerson, S. U., Kapikian, A. Z., Ticehurst, J., London, W., et al. (1993). ELISA for antibody to hepatitis E virus (HEV) based on complete open-reading frame 2 protein expressed in insect cells: Identification of HEV infection in primates. Journal of Infectious Diseases, 168(2), 369–378.PubMedGoogle Scholar
- Wang, Y., Zhang, H., Li, Z., Gu, W., Lan, H., Hao, W., et al. (2001). Detection of sporadic cases of hepatitis E virus (HEV) infection in China using immunoassays based on recombinant open reading frame 2 and 3 polypeptides from HEV genotype 4. Journal of Clinical Microbiology, 39(12), 4370–4379.PubMedCrossRefGoogle Scholar
- Widdowson, M. A., Jaspers, W. J., van der Poel, W. H., Verschoor, F., de Roda Husman, A. M., Winter, H. L., et al. (2003). Cluster of cases of acute hepatitis associated with hepatitis E virus infection acquired in the Netherlands. Clinical Infectious Diseases, 36(1), 29–33.PubMedCrossRefGoogle Scholar
- Withers, M. R., Correa, M. T., Morrow, M., Stebbins, M. E., Seriwatana, J., Webster, W. D., et al. (2002). Antibody levels to hepatitis E virus in North Carolina swine workers, non-swine workers, swine, and murids. The American Journal of Tropical Medicine & Hygiene, 66(4), 384–388.Google Scholar
- Yazaki, Y., Mizuo, H., Takahasi, M., Nishizawa, T., Sasaki, N., Goatnada, Y., et al. (2003). Sporadic acute hepatitis E in Hokkaiso, Japan, may be food-borne, as suggested by the presence of hepatitis E virus in pig liver as food. Journal of General Virology, 84(9), 2351–2357.PubMedCrossRefGoogle Scholar
- Zhang, Y., McAtee, P., Yarbourgh, P. O., Tam, A. W., & Fuerst, T. (1997). Expression, characterization, and immunoreactivities of a soluble hepatitis E virus putative capsid protein species expressed in insect cells. Clinical Diagnostic Laboratory Immunology, 4(4), 423–428.Google Scholar