Neutralization of Toscana virus is partially mediated by antibodies to the nucleocapsid protein

Abstract

The envelope glycoproteins G1/G2 of Toscana virus (TOSV) seem to have the most important protective role in stimulating antibodies against the disease in humans, as well as antibodies against the Nucleoprotein (N), a partial neutralizing activity. Mice immunized with TOSV recombinant Nucleoprotein developed a strong humoral response to the TOSV that revealed the presence of neutralizing antibody than in vitro assay. The neutralizing antibody titre of mice immunized with the whole TOSV was analyzed before and after absorption of the sera with the recombinant N protein. A decrease of the neutralizing activity was observed in the treated sera. Similar results were obtained absorbing human anti-TOSV positive sera with the recombinant N protein. This study was designed to identify the nature of antibodies produced against the N protein of TOSV in mice and to establish correlation with antibodies produced in humans by natural infection.

Performance of Various Commercial Assays for the Detection of Toscana Virus Antibodies

“…For TOSV IgG, the commercial assays evaluated in this study were observed to display slight to fair agreement with an average percentage of 57.8%. This relatively low agreement observed is presumably due to the viral antigens employed, as well as differences in assay configurations, as displayed previously (Calisher et al 1999, Cusi et al 2001, Di Bonito et al 2002. While virus-infected cells are the source of the antigens in the IIFT, recombinant N protein of TOSV is used in the ELISA and IB, although in different matrices.…”

“…This is more likely to occur when partially purified virus extracts or SFV-infected cells are employed as the source of antigens in the assay used for the diagnosis or surveillance (Schwarz et al 1995, Magurano andNicoletti 1999). During human immune response to TOSV infection, N protein has been identified as the immunodominant antigen, which has later been used for developing diagnostic assays with recombinant proteins (Ciufolini et al 1999, Cusi et al 2001. The recombinant N protein-based assays have been previously evaluated and have performed well, especially for acute TOSV infections (Valassina et al 1998, Soldateschi et al 1999).…”

“…To discriminate among related phleboviruses, it is recommended to complement recombinant N-protein-based serodiagnosis with VNT (Calisher et al 1999, Dionisio et al 2003. However, phlebovirus neutralization has been shown to be mediated only partially by antibodies to the nucleocapsid, and immunity to viral envelope glycoproteins also plays a role (Cusi et al 2001, Di Bonito et al 2002. The immune response to viral glycoproteins has been shown to differ among patients, whereas all exposed persons develop antibodies against linear epitopes of N protein (Di Bonito et al 2002).…”

Identification of a Neutralizing Epitope on TOSV Gn Glycoprotein

“…Sf9 cells were infected with baculovirus expressing the ectodomains of TOSV glycoproteins (Bac-Gn and Bac-Gc) as described elsewhere [ 22 ]. Lenti-X HEK293 were transfected with 1µg of Gn-expressing plasmid using the GeneJuice reagent (Merck-Millipore, Milan, Italy), as suggested by the manufacturer.…”

“…In several members of the family, such as Rift Valley fever virus (RVFV), La Crosse virus (LACV), Hantaan virus (HTNV), and TOSV, this neutralizing activity is a property of anti Gn-Gc antibodies [ 15 , 16 , 17 , 18 , 19 , 20 , 21 ]. In TOSV, however, a partial neutralizing activity was also shown for anti-N antibodies [ 22 ].…”