Construction of the recombinant plasmids encoding nonstructural proteins of SFTS virus and immunogenicity study

Objective To study the construction and humoral immunogenicity of the recombinant plasmids encoding nonstructural proteins(NSs) of severe fever with thrombocytopenia syndrome(SFTS) virus(SFTSV). Methods Recombinant plasmids encoding NSs gene and optimized NSs gene of SFTSV were constructed by polymerase chain reaction(PCR) and cloned into eukaryotic expression vector pJW4303, which were named pJW4303-NSs and pJW4303-NSs-opt, respectively. Then, the plasmid pJW4303-NSs was tagged with Flag, named pJW4303-NSs-Flag. Meanwhile, Nhe I restrict site and tissue plasminogen activator(tPA) signal sequence were inserted to construct bi-optimized recombinant plasmid named pJW4303-tPA-NSs-opt. All plasmids were identified by sequencing. The transient expression of NSs was confirmed by Western blotting in human embryonic kidney 293T cells. The NSs-specific IgG antibodies in BALB/c mice which were immunized by intramuscular injection with electroporation were examined by enzyme-linked immunosorbent assay(ELISA). Results The recombinant plasmids pJW4303-NSs, pJW4303-NSs-opt, pJW4303-tPA-NSs-opt and pJW4303-NSs-Flag were successfully constructed. The expression of NSs was confirmed in lysates and supernatants of 293T cells. The NSs-specific IgG responses of all three recombinant plasmids were detected by ELISA in BALB/c mice. It was found that optimized recombinant plasmid pJW4303-NSs-opt elicited higher levels of the NSs-specific IgG than that of pJW4303-NSs in week 6, 8 which induced stronger immune response. Conclusions The recombinant plasmids encoding SFTSV NSs possess the satisfied immunogenicity. In addition, the plasmid pJW4303-NSs-opt could induce the strongest humoral immune response. Key words: Severe fever with thrombocytopenia syndrome(SFTS) virus; Nonstructural protein; Recombinant plasmid; Humoral immune response