title: Bacterial expression of Crimean-Congo hemorrhagic fever virus nucleoprotein and its evaluation as a diagnostic reagent in an indirect ELISA.
abstract: Crimean-Congo hemorrhagic fever virus (CCHFV) is a tick-borne viral zoonosis distributed widely in Africa, Asia, Russia and the Balkans. The emergence and re-emergence of CCHFV emphasize the importance of increasing both human and veterinary surveillance and developing diagnostic capacity. Recombinant CCHFV nucleocapsid protein (NP) has been expressed using insect cells and mammalian cells and used as a diagnostic tool but bacterial expression has not been described previously. The S gene of CCHFV was codon optimized and the NP expressed in Escherichia coli from the synthetic gene. The protein was reacted against serum samples collected from confirmed CCHFV patients at varying intervals after the onset of illness from acute to convalescent stages using both an ELISA and a Western blot. To confirm that the protein was able to induce a humoral antibody response that could be detected using CCHFV antigen derived from live virus, mice were immunized and serum samples were tested using IF slides prepared from CCHFV infected Vero cells. The recombinant antigen was able to detect IgG antibody in acute and convalescent sera. In addition, a detectable IgG antibody response was induced in mice immunized using NP. The results suggest that proteins expressed in a bacterial system lacking post-translational modifications can be used in ELISA to detect IgG antibody against CCHFV in human sera which may be used for routine diagnosis and seroepidemiology.
authors: Department of Medical Microbiology and Virology, National Health Laboratory Services Universitas and Faculty of Health Sciences, University of the Free State, P. O. Box 339, Bloemfontein 9300, South Africa.
authors: Rudo R
authors: RR
authors: Samudzi
authors: Patricia A
authors: PA
authors: Leman
authors: Janusz T
authors: JT
authors: Paweska
authors: Robert
authors: R
authors: Swanepoel
authors: Felicity J
authors: FJ
authors: Burt
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000097:blood*; D000956:Antigens, Viral / Q000096:biosynthesis / Q000235:genetics; D015153:Blotting, Western; D002648:Child; D002675:Child, Preschool; D002522:Chlorocebus aethiops; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D003225:Hemorrhagic Fever Virus, Crimean-Congo / Q000235:genetics*; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood; D007223:Infant; D007231:Infant, Newborn; D051379:Mice; D009698:Nucleoproteins / Q000096:biosynthesis / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D014709:Vero Cells
chemical_list: D000914:Antibodies, Viral; D000956:Antigens, Viral; D007074:Immunoglobulin G; D009698:Nucleoproteins; D011994:Recombinant Proteins