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.

Fuxun Yu,Corazon C. Buerano,Hong Ma,Ferdinard Adungo,Bianli Xu,Yanhua Du,Xueyong Huang,Daisuke Hayasaka,Kouichi Morita

doi:10.1186/s12985-015-0350-0

Abstract

BackgroundSevere 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.MethodsThe 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.ResultsThe 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.ConclusionsThe 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.

Highlights

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging disease that was first reported in China in 2011
The rSFTSV-N protein was purified from the supernatant under native conditions
The identity of the rSFTSV-N protein was further confirmed by Western blot assay with mouse monoclonal antibody against histidine and the use of SFTS patient serum sample (Fig. 2)

Summary

Introduction

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. Severe fever with thrombocytopenia syndrome (SFTS), the disease caused by the virus has a major clinical presentations that include fever, thrombocytopenia, leukocytopenia, gastrointestinal symptoms, neurological symptoms, bleeding tendency, as well as less specific clinical manifestations [1, 2]. This disease has a case-fatality rate ranging from 2.5 to 30 % in different areas of endemicity [4]. It causes severe febrile illness with thrombocytopenia, leukopenia in the total blood cell count, and elevated levels of liver enzymes [16]

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