[Detection and antigenic characteristics of the recombinant nucleocapsid proteins of Lassa and Marburg viruses].

Two plasmid vectors, which allow the recombinant polypeptides of Lassa and Marburg viruses to be expressed in prokaryotic cells E. coli strain BL21 (DE3), were produced. The two recombinant polypeptides are able to bind specific antibodies. This provides an opportunity to use them as antigenic components of immunoassay diagnostic test kits.

[Characteristics of monoclonal antibodies against Lassa virus]. / Kharakteristika monoklonal'nykh antitel k nukleoproteidu virusa Lassa.

Some properties of monoclonal antibodies to the Lassa virus have been characterized. The competitive immunoenzyme analysis has revealed the presence of at least three antigens in the Lassa virus nucleoprotein.

[Conformational changes in Lassa viral proteins when exposed to antibodies and complement]. / Konformatsionnye izmeneniia v belkakh virusa Lassa pod deistviem antitel i komplementa.

Addition of the complement to the antigen in Lassa virus ELISA stimulated the sensitivity and specificity of the assay. The effect depended on the mouse Ig subclasses. ELISA plate sensitization with IgG2a and IgG2b increased the sensitivity of ELISA 8-16 times. Protein footprinting data suggest that the complement C1q component stimulates antibody-induced conformation changes of the antigen.

[Isolation and characteristics of low molecular weight recombinant peptides, representing antigenic domains in the NP protein of lymphocytic choriomeningitis virus]. / Poluchenie i kharakteristika nizkomolekuliarnykh rekombinantnykh peptidov, predstavliaiushchikh antigennye domeny belka NP virusa limfotsitarnogo khoriomeningita.

High- and low-molecular recombinant peptides of LCM virus nucleoprotein, representing individual immunodominant antigenic sites, were obtained in pJC40 expressing vector and studied in solid-phase enzyme immunoassay. 2-7% proteins resultant from total cellular synthesis are recombinant peptides. Comparative analysis of antigenic properties of recombinant peptides and native viral protein showed that recombinant peptides are virtually not inferior to native viral protein in antigenic properties.

[Production of infectious virus and the accumulation in Vero cells of Pichinde virus antigens detectable by an indirect fluorescent antibody method]. / Produktsiia infektsionnogo virusa i nakoplenie v kletkakh Vero antigenov virusa Pichinda, vyiavliaemykh nepriamym metodom fliuorestsiruiushchikh antitel.

The dynamics of accumulation of infectious Pichinde virus in the culture medium and virus-specific antigens in cells was studied in relation to multiplicity of infection in a multicycle experiment. Differences in the fluorescence pattern of Pichinde virus antigens in IFAT were found to depend on the use of acetone or formaldehyde for fixation of the infected cells.

[The detection of the Marburg virus antigen by solid-phase immunoenzyme analysis]. / Vyiavlenie antigena virusa Marburg metodom tverdofaznogo immunofermentnogo analiza.

Comparative studies of two variants of the enzyme-linked immunosorbent assay (ELISA) were carried out to determine the sensitivity of the detection of Marburg virus antigens in Vero cells. Both competitive and two-antibody ELISA variants detected as little as 5 ng of Marburg virus antigen. The Vero cell monolayer was found to produce 5-50 ng/0.05 ml of the virus-specific proteins at 6 to 8 days postinfection.

[Experimental evaluation of the pathogenicity of Lassa virus variants]. / Eksperimental'naia otsenka patogennosti variantov virusa Lassa.

Pathogenicity for guinea pigs and white mice of various Lassa virus variants: native, having had 1 passage in Vero cell culture and 4 passages in newborn white mouse brain; a virus having gone through 10 passages in Vero cells and 8 mouse brain passages (variant No. 10); a small-plaque clone derived from variant No. 10 by the method of plaque-to-plaque cloning (variant No. 11k), was studied. Both the native virus and variant No. 10 were found to be similarly pathogenic for susceptible laboratory animals, while the small-plaque variant of Lassa virus became non-fatal for guinea pigs and white mice. The decline of pathogenic properties in variant No. 11k was shown not to be associated with the presence of temperature-sensitive mutants or defective interfering particles.

[Rapid method of producing influenza A virus variants resistant to amantadine and remantadine and their primary characteristics]. / Uskorennyi metod polucheniia variantov virusa grippa A, rezistentnykh k amantadinu i remantadinu, i ikh pervichnaia kharakteristika.

A rapid method for production of influenza A virus variants resistant to the adamantane series derivatives, amantadine and remantadine, has been developed. The method consisted of two stages. In the first, the virus was subjected to one passage in the presence of the preparations under a liquid overlayer in a one-cycle experiment. In the second stage, the resulting virus was titrated by the plaque method, the agar overlay containing the preparations in a concentration which was not toxic for the cells. Production of large and small plaques in the presence of the preparations in agar was an indication for selection of resistant virus variants and their further study. Cross-resistance of amantadine- and remantadine-resistant variants to remantadine and amantadine, respectively, was studied. No complete cross-resistance in these viruses could be demonstrated. The amantadine-resistant virus was not inhibited by remantadine, whereas the remantadine-resistant virus was significantly inhibited by amantadine as was demonstrated by both virological methods and by induction of RNA-dependent RNA polymerase and synthesis of viral proteins. The experimental results suggest that the mechanisms of formation of influenza A virus resistance to amantadine and remantadine are not absolutely identical.

[The immunogenic activity of the Lassa virus structural proteins]. / Immunogennaia aktivnost' strukturnykh belkov virusa Lassa.

The immunogenic properties of Lassa virus GP1, GP2, and NP polypeptides were studied in rabbits. Lassa virus NP polypeptide, in contrast to GP1 and GP2 polypeptides, was shown to induce the highest titres of antibodies determined by IFA and ELISA tests. Moreover, the antibody relative avidity experiment showed that the anti-NP antibodies has ELISA index of 0.63 whereas anti-GP1 and anti-GP2 antisera had those of 0.49 and 0.28, respectively.

[Inhibition of the accumulation of the Lassa virus in Vero cells by immune gamma globulin and complement]. / Podavlenie nakopleniia virusa Lassa v kletkakh Vero immunnym gamma-globulinom i komplementom.

Specific inhibition of Lassa virus replication in Vero cells was found to be better achieved with immune gamma globulin in combination with complement than with gamma globulin alone. According to the authors, the inhibitory effect of these preparations is due to the cyto-destructive action of antibodies and complement on the infected cells.

[False-positive reactions in laboratory diagnosis of Lassa, Marburg, and Ebola viral hemorrhagic fevers and AIDS]. / Lozhnopolozhitel'nye reaktsii pri laboratornoi diagnostike virusnykh gemorragicheskikh likhoradok Lassa, Marburg, Ebola i SPIDa.

Sera of normal subjects and AIDS patients living in Minsk and Odessa were tested for antibodies to hazardous viral infections Lassa, Marburg, and Ebola. Four to 16% of examinees were seropositive to Ebola virus, 0.8 to 2.3% to Lassa, and up to 0.8% to Marburg virus. Common B-epitopes were found in viruses belonging to different families: Lassa, Ebola, and HIV. Antibodies specific to these viruses antigens were found in the reference sera to influenza A and B, respiratory syncytial virus, and adenovirus. Sera of convalescents after malaria and of AIDS patients contained antibodies to Lassa virus.

[Monoclonal antibodies to the Machupo virus: their isolation and preliminary characteristics]. / Monoklonal'nye antitela k virusu Machupo: poluchenie i predvaritel'naia kharakteristika.

Six monoclonal antibody-producing hybridoma cell lines were generated by fusion of NS-1 myeloma cells with BALB/c immune splenocytes. Monoclonal antibodies (MCA) specific to Machupo virus NP protein were used to study cross-reactivity between pathogenic and nonpathogenic arenaviruses. It was shown that 3140 MCA cross-reacted in IFA with Lassa, Tacaribe, and Tamiami arenaviruses whereas 3101 MCA reacted with Machupo virus alone. It was assumed that these 3101 MCA could be used for differentiation of Machupo virus in IFA.

[The isolation and characteristics of recombinant monoclonal antibodies to the Lassa virus]. / Poluchenie i kharakteristika rekombinantnykh monoklonal'nykh antitel k virusu Lassa.

Recombinant monoclonal antibodies to Lassa virus were produced. The reactivity of the monoclonal antibodies was studied by indirect fluorescence antibody (IFA) technique, enzyme-linked immunosorbent assay (ELISA), and radioimmunoprecipitation method. The observed reactivity did not correlate with IgG isotyping groups.

[Recombinant monoclonal antibodies to the Lassa virus: the formation of reactive paratopes]. / Rekombinantnye monoklonal'nye antitela k virusu Lassa: formirovanie reaktivnykh paratopov.

Eighteen hybrid lines secreting recombinant monoclonal antibodies to Lassa virus were produced by fusion of mouse splenocytes with antibody-secreting X-63 myeloma cells. Interrelations between the structure and reactivity of the antibodies were studied by different serological and immunochemical methods. Monoclonal antibodies were divided into different groups according to their serological properties and macromolecular structure. A comparative analysis of the structure and reactivity of the recombinant monoclonal antibodies showed that the light and heavy Ig-specific chains could form the reactive antibodies when the chains were present in different paratopes of Ig molecules.

[Serological evidence of Lassa virus circulation in the Republic of Guinea]. / Serologicheskie dokazatel'stva tsirkuliatsii virusa Lassa v Gvineiskoi Respublike.

Seroepidemiological investigations in Guinea were carried out to estimate the areas of Lassa virus circulation. The recombinant protein of Lassa virus nucleocapsid was used as the antigen to analyse blood sera by ELISA. In some regions, from 30 to 54.9% of the population had antibodies to Lassa virus, but in others only 6-7%.

[The localization and preliminary characteristics of antigenic sites (B epitopes) in the nucleocapsid protein of the Lassa virus]. / Lokalizatsiia i predvaritel'naia kharakteristika antigennykh saitov (B-épitopov) v belke nukleokapsida virusa Lassa.

Nucleocapsid protein of Lassa virus contains 4 epitope sites. Three of them were localized in amino acid position 123-1 27, 337-346, and 518-527. Monoclonal antibody 1108 specific to Lassa virus NP-protein reacted with 123-127 synthetic peptide in solid-phase ELISA and precipitated nucleocapsid proteins of Machupo, Tacaribe, and Mopeia arenaviruses in RIP.

[Virus-specific proteins and RNA of the virus of hemorrhagic fever with renal syndrome]. / Virusspetsificheskie belki i RNK virusa gemorragicheskoi lokhoradki s pochechnym sindromom.

Virus-specific proteins G1, G2, and N with molecular weights of 70, 55-57, and 50 kilodaltons, respectively, were detected by radioimmunodiffusion tests in VERO E-6 cells infected with strains of virus of hemorrhagic fever with renal syndrome (HFRS) isolated in the European USSR from a patient with HFRS, a fatal human case of HFRS (the strains K-27 and P-360) and from a bank vole (strain CG-1820). The sera from human convalescents after HFRS in the European USSR and rat sera prepared with the CG-1820 strain precipitated proteins possessing similar electrophoretic characteristics from a lysate of cells infected with the CG-1820, K-27 and P-360 strains. The sera from human HFRS convalescents in the Far East did not precipitate protein Gl. The viral RNA derived by immunosorption method from intracellular nucleocapsids of CG-1820 strain and strain 4590 (isolated from Ap. Peninsulae in the Far East) contained 3 classes of molecules: L, M, and S. L- and M-RNA of these strains had the same molecular weight (1.62 and 1.38 megadaltons). The molecular weight of S-RNA of the strain 4590 was 0.76 megadalton and that of the CG-1820 strain 0.83 megadalton. It is assumed that there are species differences among the viruses, causative agents of HFRS, circulating in the European and Far East regions of the USSR.

[The production of antibody-producing hybridomas to the Lassa virus]. / Poluchenie antiteloprodutsiruiushchikh gibridom k virusu Lassa.

The work deals with obtaining hybrid cell lines producing monoclonal antibodies to Lassa arenavirus. To obtain preparations for the screening of hybridomas by indirect immunofluorescence techniques, the dynamics of the accumulation of Lassa virus antigen in cell cultures Vero and 4647 was studied. The maximum accumulation of the virus antigen in Vero cells was shown to occur on day 3 after inoculation with a dose of 1.0 PFU/ml. The influence of different doses of gamma radiation on the infectious and antigenic activity of the virus was studied. The expediency of using a dose of 20.0 kGy for the irradiation of the virus was shown. The optimum schedule for the immunization of BALB/c mice was worked out, which made it possible to obtain activated mouse spleen cells used for the construction of hybridomas. The capacity of hybrid cells, injected into syngeneic mice, for the generation of tumors was shown.

[The dynamics of the formation of IgG subclasses in Balb/c and CBA mice infected with the LCM, Lassa and Mopeia viruses and their role in diagnosis]. / Dinamika obazovaniia podklassov IgG u myshei Balb/c i CBA pri zarazhenii virusami LKhM, Lassa, Mopeiia i ikh rol' v diagnostike.

The dynamics of the induction of individual IgG subclasses in BALB/c and CBA mice and their role on the diagnostics of arenaviruses LCM, Lassa and Mopeia was studied. The study demonstrated that in solid-phase enzyme immunoassay (EIA) isotypes IgG2a and IgG2b to virus Mopeia could be used for the differentiation of virus Mopeia from viruses LCM and Lassa, and the antigen of virus Mopeia could be used for the preparation of diagnostic EIA systems not only to nonpathogenic virus Mopeia, but also to pathogenic viruses LCM and Lassa.

[Effect of amantadine (1-aminoadamantane) and remantadine (alpha-methyl-1-adamantane methyl amine) hydrochlorides on formation of influenza A viral proteins]. / Sravnitel'noe izuchenie deistviia solianokislykh amantadina (1-aminoadamantana) i remantadina (al'fa-metil-1-adamantanmetilamina) na obrazovanie belkov virusa grippa A.

Inhibitory effect of amantadine (I-amino adamantane) and remantadine (alpha-methyl-1-adamantane methylamine) hydrochlorides on reproduction of influenza A virus as well as, in particular, on formation of virus specific proteins was studied in cell culture. These drugs were dissimilar; they resembled each other in the patients of their effect as a function of the course of administration into the cells infected with the virus. Both preparations inhibited the synthesis of virus specific proteins exhibiting distinct effects under conditions of preliminary or immediate administration following the infection. At the same time, the inhibitory properties of remantadine were maintained at the subsequent period after infection; these were not observed in the amantadine treatment.