title: Application of recombinant severe fever with thrombocytopenia syndrome virus nucleocapsid protein for the detection of SFTSV-specific human IgG and IgM antibodies by indirect ELISA.
abstract: BACKGROUND
  Severe fever with thrombocytopenia syndrome (SFTS) is an emerging disease that was first reported in China in 2011. It is caused by SFTS virus (SFTSV) which is a member of the Phlebovirus genus in the Bunyaviridae family. SFTSV has been classified as a BSL3 pathogen. There is a need to develop safe and affordable serodiagnostic methods for proper clinical management of infected patients.
  
  
  METHODS
  The full length nucleocapsid (N) gene of SFTSV Yamaguchi strain was amplified by RT-PCR and cloned to an expression vector pQE30. The recombinant (r) SFTSV-N protein was expressed by using Escherichia coli (E. coli) expression system and purified under native conditions. rSFTSV-N protein based indirect IgG and IgM enzyme linked immunosorbent assay (ELISA) systems were established to detect specific human IgG and IgM antibodies, respectively. One hundred fifteen serum samples from clinically suspected-SFTS patients were used to evaluate the newly established systems and the results were compared with the total antibody detecting sandwich ELISA system.
  
  
  RESULTS
  The native form of recombinant (r) SFTSV-N protein was expressed and purified. Application of the rSFTSV-N protein based indirect IgG ELISA to the 115 serum samples showed results that perfectly matched those of the total antibody sandwich ELISA with a sensitivity and specificity of 100 %. The rSFTSV-N protein based indirect IgM ELISA missed 8 positive samples that were detected by the total antibody sandwich ELISA. The sensitivity and specificity of rSFTSV-N-IgM capture ELISA were 90.59 and 100 %, respectively.
  
  
  CONCLUSIONS
  The rSFTSV-N protein is highly immunoreactive and a good target for use as an assay antigen in laboratory diagnosis. Its preparation is simpler in comparison with that used for the total antibody sandwich system. Our rSFTSV-N protein-based IgG and IgM ELISA systems have the advantage of distinguishing two types of antibodies and require small volume of serum sample only. They are safe to use for diagnosis of SFTS virus infection and especially fit in large-scale epidemiological investigations.
authors.affiliation: Department of Virology, Institute of Tropical Medicine, Nagasaki University, 1-12- 4, Sakamoto, Nagasaki, 852-8523, Japan. yufuxun1@nagasaki-u.ac.jp.
authors.forename: Fuxun
authors.initials: F
authors.lastname: Yu
authors.affiliation: Henan Center for Disease Control and Prevention, Zhengzhou, China. duyh325@163.com.
authors.forename: Yanhua
authors.initials: Y
authors.lastname: Du
authors.affiliation: Henan Center for Disease Control and Prevention, Zhengzhou, China. hxyzzu@163.com.
authors.forename: Xueyong
authors.initials: X
authors.lastname: Huang
authors.affiliation: Henan Center for Disease Control and Prevention, Zhengzhou, China. mahong@hncdc.com.cn.
authors.forename: Hong
authors.initials: H
authors.lastname: Ma
authors.affiliation: Henan Center for Disease Control and Prevention, Zhengzhou, China. bianlixu@163.com.
authors.forename: Bianli
authors.initials: B
authors.lastname: Xu
authors.affiliation: Department of Virology, Institute of Tropical Medicine, Nagasaki University, 1-12- 4, Sakamoto, Nagasaki, 852-8523, Japan. ferdinard.adungo@gmail.com.
authors.forename: Ferdinard
authors.initials: F
authors.lastname: Adungo
authors.affiliation: Department of Virology, Institute of Tropical Medicine, Nagasaki University, 1-12- 4, Sakamoto, Nagasaki, 852-8523, Japan. hayasaka@nagasaki-u.ac.jp.
authors.forename: Daisuke
authors.initials: D
authors.lastname: Hayasaka
authors.affiliation: Department of Virology, Institute of Tropical Medicine, Nagasaki University, 1-12- 4, Sakamoto, Nagasaki, 852-8523, Japan. ccbuerano@hotmail.com.
authors.forename: Corazon C
authors.initials: CC
authors.lastname: Buerano
authors.affiliation: Department of Virology, Institute of Tropical Medicine, Nagasaki University, 1-12- 4, Sakamoto, Nagasaki, 852-8523, Japan. moritak@nagasaki-u.ac.jp.
authors.forename: Kouichi
authors.initials: K
authors.lastname: Morita
mesh_terms: D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D015870:Gene Expression; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood / Q000276:immunology*; D007075:Immunoglobulin M / Q000097:blood / Q000276:immunology*; D019590:Nucleocapsid Proteins / Q000235:genetics* / Q000276:immunology* / Q000302:isolation & purification; D010217:Phlebotomus Fever / Q000175:diagnosis / Q000276:immunology* / Q000821:virology; D016856:Phlebovirus / Q000235:genetics* / Q000276:immunology*; D011994:Recombinant Proteins
chemical_list: D007074:Immunoglobulin G; D007075:Immunoglobulin M; D019590:Nucleocapsid Proteins; D011994:Recombinant Proteins