Founded in 1975 as the all Union Research Institute of Applied Enzimology, currently,  the Institute of Biotechnology is mainly involved in research and training in the fields of biotechnology and molecular biology, including research and development of recombinant biomedical proteins, genetic and molecular studies of restriction modification phenomenon, developing of viruses diagnostics, epigenetic study of small RNA, drug design and synthesis, bioinformatics.

Hantavirus vaccine and diagnostic tools

Spausdinti

Linas Antoniukas, Rita Vorobjovienė, Rasa Petraitytė, Aušra Ražanskienė

The project was supported by grant Nr. T-83/07 from Lithuanian State Science and Studies Foundation.

Hantaviruses represent a separate genus Hantavirus of the family Bunyaviridae. They contain a tripartite RNA genome of negative polarity. The genome segments M (medium) and L (large) encode a glycoprotein precursor which is co-translationally processed into G1 and G2, and an RNA-dependent RNA polymerase. The S (small) genome segment codes for the nucleocapsid (N) protein. We have described the high-level yeast expression of authentic and amino-terminally His-tagged rN protein of PUUV (strain Vranica/Hällnäs) which are able to induce a protective immune response in bank voles, the natural host of PUUV (Dargevičiūtė et al., 2002).

High-level expression in yeast and purification of high yields of rN proteins of hantavirus species originating from Asia (HTNV) and different regions of Europe (PUUV strains Sotkamo and Kazan from Finland and Russia respectively; DOBV-Slk from Slovakia and DOBV-Slo from Slovenia) was also performed. The rN proteins were characterized in terms of stability, nucleic acid and endotoxin contamination, antigenicity and immunogenicity (Razanskiene et al., 2004).

Hantavirus seroprevalence was investigated in Lithuania.

Recently we started to investigate Hantavirus distribution in Lithuanian population (Sandmann et al., 2005). This work was extended with patients from dialysis centers. Serums of 218 patients from four dialysis centers of Kaunas district were tested for hantavirus specific antibodies by using the IgG antibody-capture ELISA. Antibodies against Dobrava/Hantaan and Puumala hantaviruses were found in 16 patients (seroprevalence 7.4%). Most of the sera were positive for Dobrava/Hantaan hantavirus (81%). Seroprevalence was significantly higher in older patients. In general the seroprevalence of dialysis patients is similar to previously estimated seroprevalence of patients from oncology center (Dargevicius et al. 2007).

New methods for hantavirus diagnostics

For the serological detection of human infections by hantaviruses relevant for Europe, we developed monoclonal antibody capture immunoglobulin G (IgG) and IgA enzyme-linked immunosorbent assays (ELISAs) based on yeast-expressed nucleocapsid proteins of Puumala and Dobrava hantaviruses. The specificities of the Puumala and Dobrava virus-specific IgA, and IgG ELISAs were found to be 100%. The sensitivities of these ELISAs were determined to be 100% with panels of characterized anti-Puumala or anti-Dobrava virus-positive human serum samples. In most cases, Puumala and Dobrava virus infections could be differentiated by ELISA reactivity alone, i.e., endpoint titration with homologous and heterologous antigens. (Meisel et.al. 2006)

Indirect and capture enzyme-linked immunosorbent assays (ELISAs) for detection of Hantaan virus (HTNV)-specific immunoglobulins G (IgG) and M (IgM) in human serum samples were developed on the basis of recombinant yeast-expressed nucleocapsid (N) protein of HTNV. The sensitivity of the indirect IgG and IgM ELISA tests was both 100% and the specificity of the indirect IgM and IgG ELISA test was 98% and 99%, respectively. The sensitivity and specificity of the capture IgM ELISA was 100% and 97%, respectively. The novel assays were found to detect HTNV-specific antibodies in acute phase sera from suspected HFRS patients in China. The results indicate that these novel ELISAs are suitable for the diagnosis of HTNV and for sero-epidemiological studies (Petraityte et al., 2007a).

For the detection of Hantaan virus in the oral fluid of humans, we have developed a monoclonal antibody-based capture enzyme-linked immunosorbent IgM assay (IgM capture ELISA) and indirect enzyme-linked immunosorbent IgG and IgM assays (indirect IgG and IgM ELISAs) for paired serum and oral fluid samples using the Saccharomyces cerevisiae yeast-expressed nucleocapsid protein of the Hantaan-Fojnica virus. The sensitivity and specificity of the oral fluid IgM capture ELISA in comparison with the results of the serum Hantaan virus IgM assay were 96.7% and of 94.9%, respectively. In conclusion, the IgM capture ELISA can be used with oral fluid instead of serum samples for the diagnosis of Hantaan virus infection (Petraityte et al., 2007b).