[Functional activity of glycoproteins from interepidemic isolates of the influenza virus]. / Izuchenie funktsional'noi aktivnosti glikoproteidov mezhépidemicheskikh izoliatov virusa grippa.

The results of the study into the function of glycoproteins of influenza viruses isolated in the 1987 interepidemic period. Natural isolates were found to have virus particles with deficient neuraminidase activity but retained hemolytic and infectious activity. Biological sequences of the lack of neuraminidase activity in the isolates consisted in disorders of sialic acid metabolism in the infected cells, blocking of the receptor site of hemagglutinin and increased sensitivity to inhibitors. It is assumed that the interepidemic virus isolates represent a biologically heterogeneous population in which particles with deficient neuraminidase activity are prevalent.

[Antigenic specificity of neuraminidase and structure of neuraminidase gene of influenza A(H1N1) serotype Hsw1N1 viruses isolated from man and animals]. / Antigennaia spetsifichnost' neiraminidazy i struktura genov neiraminidazy virusov grippa A(H1N1)-serovariant Hsw1N1, vydelennykh ot cheloveka i zhivotnykh.

The enzyme immunoassay and neuraminidase activity inhibition test using polyclonal and monospecific antineuraminidase sera were employed to establish the similarities and differences in the antigenic structure of neuraminidase of influenza A viruses (H1N1), serovariant Hsw1N1, isolated from man in Alma-Ata (USSR), 1984, New Jersey (USA), 1976, and Pazardjik (BPR), 1982, as well as from swine and birds. Oligonucleotide mapping revealed significant structural differences in the genes coding for neuraminidase of Hsw1N1 viruses. The experimental results indicate a high degree of the enzyme variability in this group of viruses.

[Isolation and characteristics of cloned variants of influenza virus A/USSR/13/81 (H1N1-N3)]. / Poluchenie i kharakteristika klonirovannykh variantov virusa grippa A/SSSR/13/81 (H1N1-N3).

Cloning in chick embryos and MDCK cell culture of influenza A/USSR/13/81 (H1N1-N3) virus isolated during virological examinations of autopsy materials from a child who had died from acute respiratory virus infection yielded three subpopulations of clones differing in antigenic, biological, physico-chemical properties and glycoprotein structures. One subpopulation contained hemagglutinin (HA) similar to that of the A/PR/8/34 strain and neuraminidase (NA) N3, the other HA similar to that of A/WS/33 and NA N1, and the third HA of the isolate proper and NA of the both serosubtypes mentioned.

Controlled organization of multimolecular complexes of enveloped virus glycoproteins: study of immunogenicity.

Some technological and immunological problems facing the preparation of subunit viral vaccines are discussed. Solubilization of enveloped virus glycoproteins with various detergents has been studied. It has been demonstrated that a novel non-ionic detergent, MESK, can be used to prepare the glycoproteins of enveloped viruses in defined supramolecular forms: monomers, micelles, liposomes and multimeric complexes. These preparations have been tested for immunogenicity. It has been shown that the immunogenicity of glycoproteins in micellar form or in liposomes is comparable with that of the whole virus. The immunogenicity of the glycoprotein complex with the glycoside Quil A appeared to be significantly higher in comparison with the whole virus and was similar to the immunogenicity of glycoproteins mixed with Freund's complete adjuvant.

[Virologic and seroepidemiologic analyses of the circulation of influenza virus A(H1N1), similar to serovariant A(Hsw1N1), in 1984-1985 in Alma-Ata]. / Virusologicheskii i seroépidemiologicheskii analizy tsirkuliatsii virusa grippa A(H1N1), podobnogo serovariantu A(Hsw1N1), v 1984-1985 gg. v Alma-Ate.

The study made with the use of virological, seroepidemiological, radioimmunological and immunological techniques revealed that influenza virus A, antigenically similar to influenza virus A/swine/Iowa/15/30, circulated in Alma-Ata in 1984-1985. The role of these viruses in the seasonal rise of influenza morbidity at the end of 1984 was established. From nasal washings and blood clots obtained from patients, as well as from dissection material, 12 strains were isolated. These strains were similar to serovariant A/swine/owa/15/30 and differed from influenza virus A/New Jersey/76.

[Inhibition of the infectivity of enveloped viruses while retaining the specific biological activity of the viral proteins]. / Podavlenie infektsionnosti obolochechnykh virusov s sokhraneniem spetsificheskoi biologicheskoi aktivnosti virusnykh belkov.

A method of inactivation of enveloped viruses by treatment of virus-containing material with a nonionic detergent MESK was studied. Treatment with the detergent of allantoic or culture fluids containing influenza, parainfluenza, herpes simplex, and Venezuelan equine encephalomyelitis viruses was shown to result in significant (to 10 lg ID50) decrease of the infectivity of the viruses. At that, the specific biological activity of viral proteins: the hemagglutinating and neuraminidase activities in the case of influenza and parainfluenza viruses, and the hemagglutinating activity in the case of Venezuelan equine encephalomyelitis virus, remained unchanged. Treatment of the viruses with the detergent in the presence of 2% BSA, 5% gamma-globulin, or 50% blood serum also inactivated the infectivity of virions. The inactivating effect of the MESK detergent is associated with solubilization of external glycoproteins of the virus envelope. The method of mild nondenaturating inactivation of enveloped viruses with the MESK detergent may prove to be useful in laboratory practice.

[Solubilization of glycoproteins from enveloped viruses by detergents]. / Soliubilizatsiia glikoproteidov obolochechnykh virusov detergentami.

The effects of some known ionic and nonionic detergents as well as that of a novel nonionic detergent MESK on various enveloped viruses were investigated. It was found that nonionic detergens (MESK, Triton X-100, octyl-beta-D-glucopyranoside) selectively solubilize glycoproteins of enveloped viruses. The most mild selective action is exerted by the nonionic detergent MESK. Using this detergent, pure preparations of glycoproteins of influenza, parainfluenza, equine Venezuela encephalomyelitis, rabies, vesicular stomatitis and herpes viruses were obtained. The procedure of isolation of purified glycoproteins includes incubation of viral suspensions with MESK, removal of subviral structures by centrifugation and purification of glycoproteins from detergent admixtures by dialysis. Purified glycoproteins retain their native structure and a high biological activity and immunogenicity. MESK seems to be due a perspective tool in the production of subunit vaccines.

[Isolation of glycoproteins of the Venezuelan equine encephalomyelitis virus and an evaluation of their immunogenic activity]. / Vydelenie glikoproteidov virusa venesuél'skogo éntsefalomielita loshadei i otsenka ikh immunogennoi aktivnosti.

Isolation of purified Venezuelan equine encephalomyelitis virus glycoproteins by means of a new Soviet nonionic detergent, MESK, is described. The MESK detergent was shown to permit isolation of approximately 70% of virus particle glycoproteins. The resulting preparation had a high hemagglutinating activity, contained no admixtures of foreign proteins and was not infectious. The study of the immunogenicity of purified glycoproteins in experimental mice and rabbits showed them to be capable of inducing high levels of serum antibodies. The immunogenicity of the isolated glycoproteins was comparable to their immunogenicity as components of virus particles. Treatment of the virus with MESK detergent also yielded preparations with predominant content of capsid protein. The described procedure for disintegration and purification of viral proteins is technologically simple and may be the basis for manufacture of a subunit vaccine. The resulting material may also be used for preparation of diagnosticums and in laboratory studies.

[Immunogenicity of a subunit influenza vaccine in experiments on animals]. / Izuchenie immunogennosti sub"edinichnoi grippoznoi vaktsiny v opytakh na zhivotnykh.

The data on isolation of purified influenza virus glycoproteins and their reconstruction into liposomes by means of a new nonionic detergent, MESK, are presented. Sedimentational and flotational distribution of glycoprotein and liposome preparations was studied. In the presence of the detergent, the isolated glycoproteins were shown to occur mostly in a monomeric form. Removal of the detergent by dialysis resulted in formation of protein micelles, and the presence of exogenously introduced lipids in reconstruction of liposomes heterogeneous in composition. The resulting preparations had a high degree of biochemical purity and biological activity meeting the requirements for subunit vaccine properties. The immunogenic potency of a subunit influenza vaccine produced with the use of the MESK detergent was studied. The glycoproteins isolated with the use of MESK were shown to be comparable in their immunogenic potency with glycoproteins comprising viral particles. The influence of the form of glycoproteins presentation on their immunogenic potency was studied. Glycoproteins in the form of micelles and as components of liposomes were found to have a good immunogenic potency and to be able to induce protective immunity preventing experimental influenza infection in mice. Monomeric forms of glycoproteins had no such properties.

Isolation and studies of myxovirus glycoproteins.

The isolation of ortho- and paramyxovirus glycoproteins using a new nonionic detergent (MESK) is reported. MESK was shown to solubilize most of the viral envelope glycoproteins without decreasing their biologic activity. Solubilized glycoproteins are not contaminated by any internal viral proteins or by appreciable quantities of viral envelope lipids. The removal of MESK by dialysis resulted in the formation of glycoprotein micelles. The immunogenic activity of isolated glycoproteins was compared to that of virus particles. Immunization with isolated glycoproteins was shown to protect mice against a lethal influenza infection. Virions were treated with MESK in the presence of exogenous egg phosphatidylcholine, detergent was removed by dialysis and the glycoprotein was reconstituted in the vesicles. This reconstitution was accompanied by restoration of the haemolytic activity of Sendai virus proteins up to that of native virus particles. The level of activity, also the morphology and buoyant density of the vesicle were dependent on the protein/lipid ratio. MESK proved to be of value for the selective solubilization of the surface glycoproteins of animal enveloped viruses and their reconstitution in liposomes.

[Effect of influenza virus glycoproteins on the permeability of pulmonary capillaries in mice]. / Vliianie glikoproteidov virusa grippa na sostoianie pronitsaemosti legochnykh kapilliarov myshei.

Isolated glycoproteins of influenza A/PR/8/34 (H1N1) administered intranasally to white mice were found to increase permeability of lung capillaries.

[Isolation of purified glycoproteins of influenza virus and analysis of their antigenic and immunogenic activity]. / Poluchenie ochishchennykh glikoproteidov virusa grippa i issledovanie ikh antigennoi i immunogennoi aktivnosti.

Investigations carried out by the authors have demonstrated that a new specially synthesized nonionic detergent, MECK, can be successfully used for obtaining pure influenza virus surface antigens in preparative amounts. Hemagglutinin and neuraminidase isolated by means of MECK retain their structure and biological activity. Special attention has been devoted to the study of the immunological characteristics of the preparations thus obtained. The high immunogenicity of solubilized hemagglutinin and the complete retention of its antigenic activity have been confirmed. The simplicity of the method for the isolation of influenza virus surface antigens and their purification from internal viral proteins and the detergent make it possible to recommend the above method for the preparative isolation of external antigens.

[Effective method of isolating influenza virus on chick embryos]. / Effektivnyi metod vydeleniia virusa grippa na kurinykh émbrionakh

An effective method for isolation of influenza virus from the nasopharyngeal specimens of patients is described consisting of interruption of the virus reproduction cycle in chorioallantoic membranes of chick embryos (CECAM) at early stages of development of the infection followed by inoculation of infected CECAM homogenate into a fresh homologous medium. The new method of virus isolation is superior to the conventional one by a higher isolation rate, economy of material and time saving which is important for identification of influenza outbreaks.