RESUMEN
A process flow diagram was elaborated for production and control of tissue culture inactivated vaccine (TCIV) against Japanese encephalitis (JE). The vaccine was prepared on the basis of the earlier patented JE virus strain (K3). Experimental laboratory JE TCIV series were obtained; their safety and high immunogenicity were tested on animals. Regulations (an instruction) for preparing and controlling JE TCIV have been worked out, which have been approved by the Academic Council of the D. I. Ivanovsky Institute of Virology.
Asunto(s)
Virus de la Encefalitis Japonesa (Especie)/inmunología , Encefalitis Japonesa/inmunología , Encefalitis Japonesa/prevención & control , Inmunización , Vacunas contra la Encefalitis Japonesa/inmunología , Vacunas contra la Encefalitis Japonesa/aislamiento & purificación , Vacunas contra la Encefalitis Japonesa/toxicidad , Animales , Anticuerpos Antivirales/sangre , Anticuerpos Antivirales/inmunología , Células Cultivadas , Embrión de Pollo , Encefalitis Japonesa/sangre , Formaldehído , Cobayas , Pruebas de Inhibición de Hemaglutinación , Esquemas de Inmunización , Inyecciones Intramusculares , Inyecciones Intraperitoneales , Vacunas contra la Encefalitis Japonesa/administración & dosificación , Ratones , Pruebas de Neutralización , ConejosRESUMEN
Genetic analysis of strains and variants of Japanese encephalitis virus from the collection of D. I. Ivanovsky Institute of Virology revealed marked differences in the complex of genetic markers, which makes possible more precise selection of the most interesting clones.
Asunto(s)
Virus de la Encefalitis Japonesa (Especie)/genética , Virus de la Encefalitis Japonesa (Especie)/clasificación , Humanos , Federación de RusiaRESUMEN
Analysis of 10 strains and 7 variants of Japanese encephalitis virus from the collection of the D. I. Ivanovskii Institute of Virology of the Russian Academy of Medical Sciences revealed marked differences between the populations in the complex of biological properties (pathogenicity, reproductive potential, antigenic activity, etc.). Of the Japanese encephalitis virus variants included in the collection attenuated clones 33 and 43 are practically the most interesting as potential candidates for diagnostic and prophylactic preparations.
Asunto(s)
Virus de la Encefalitis Japonesa (Especie)/fisiología , Variación Genética , Academias e Institutos , Animales , Virus de la Encefalitis Japonesa (Especie)/genética , Virus de la Encefalitis Japonesa (Especie)/aislamiento & purificación , Virus de la Encefalitis Japonesa (Especie)/patogenicidad , Variación Genética/genética , Ratones , Ratas , Federación de Rusia , Pase Seriado , Virología , Cultivo de Virus , Replicación ViralRESUMEN
Experiments with attenuated clones of Venezuelan equine encephalomyelitis virus and Eastern equine encephalomyelitis virus were carried out to study the regularities in changes of biological properties of virus in "undiluted" passages and in passages by subcultivation of small doses. In the latter case the biological activity of the virus remained at the initial low level but in "undiluted" passages it increased, due to accumulation in the population of clones with altered plaque phenotype and increased reproductive potential. In a number of cases this virus had a higher level of residual virulence than the original one. The evidence that the main source of virus variability in the "undiluted" passages lies, in genetic interaction in which defective virus takes part, is presented. Mixed infection with participation of defective interfering particles and the genetic interaction occurring in it are considered to be the mechanism which is conducive to restoration of biological activity of alphaviruses.
Asunto(s)
Alphavirus/fisiología , Alphavirus/genética , Alphavirus/patogenicidad , Animales , Embrión de Pollo , Virus Defectuosos/genética , Virus de la Encefalitis Equina del Este/genética , Virus de la Encefalitis Equina del Este/patogenicidad , Virus de la Encefalitis Equina del Este/fisiología , Virus de la Encefalitis Equina Venezolana/genética , Virus de la Encefalitis Equina Venezolana/patogenicidad , Virus de la Encefalitis Equina Venezolana/fisiología , Ratones , Fenotipo , Interferencia Viral , Ensayo de Placa Viral , Virulencia , Cultivo de Virus/métodos , Replicación ViralRESUMEN
Some genetic and biological properties of the multiploid -forming mutant of Venezuelan equine encephalomyelitis virus as well as of standard and multiploid virions isolated from its population were analysed. The mutant steadily retained the capacity for formation of multiploid virions comprising up to 20% of the population both in passages in CEF culture and in the progeny of multiploid and standard virions. Owing to this, the concentration of multiploid virions in the population of this mutant remains at a constant level. Formation of multiploid virions compensates for the temperature-sensitive defect inherent in the standard virions which is associated with later stages of virus replication.
Asunto(s)
Virus de la Encefalitis Equina Venezolana/fisiología , Mutación , Poliploidía , Animales , Embrión de Pollo , Virus de la Encefalitis Equina Venezolana/genética , Virus de la Encefalitis Equina Venezolana/aislamiento & purificación , Fenotipo , Temperatura , Ensayo de Placa Viral , Virión/genética , Virión/aislamiento & purificación , Virión/fisiología , Cultivo de VirusRESUMEN
A natural process of infection with multiploid virions was stimulated and the genetic effects occurring in this infection were studied in experiments with homogeneous attenuated strain of Venezuelan equine encephalomyelitis virus producing the smallest plaques. The process of infection with multiploid virions was reproduced by infecting the cells with artificially obtained aggregates. Under these conditions, small- and large-plaque virus was produced which in some cases had a higher virulence than the parental strain. A possible mechanism of this phenomenon and its practical implications for the evaluation of the properties of attenuated alphavirus strains is discussed.
Asunto(s)
Alphavirus/genética , Variación Genética , Infecciones por Togaviridae/microbiología , Virión/genética , Alphavirus/patogenicidad , Animales , Embrión de Pollo , Virus de la Encefalitis Equina Venezolana/genética , Virus de la Encefalitis Equina Venezolana/patogenicidad , Encefalomielitis Equina Venezolana/microbiología , Genes Virales , Ensayo de Placa Viral , Virión/patogenicidad , Virulencia , Cultivo de VirusRESUMEN
The main molecular biology parameters of an attenuated mutant DMS-20/6 of eastern equine encephalomyelitis virus derived by treatment with dimethylsulphate of the wild type virus (strain No. 627) were determined. The sedimentation coefficient of sucrose density gradient purified and concentrated virus was 280 S, the buoyant density of virions in sucrose density gradient was 1.19 g/cm3. The DMS-20/6 virion had 3 proteins with molecular weights of 56, 50, and 34 kilodaltons, and the size of virions by negative staining was 58-77 nm.
Asunto(s)
Alphavirus/análisis , Virus de la Encefalitis Equina del Este/análisis , Mutación , Centrifugación Zonal , Dimetilsulfóxido/farmacología , Electroforesis en Gel de Poliacrilamida , Virus de la Encefalitis Equina del Este/efectos de los fármacos , Virus de la Encefalitis Equina del Este/aislamiento & purificación , Microscopía Electrónica , Proteínas Virales/análisis , Proteínas Virales/aislamiento & purificación , Virión/análisis , Virión/efectos de los fármacos , Virión/aislamiento & purificación , Virulencia/efectos de los fármacosRESUMEN
Comparative data with regard to the properties of Venezuelan equine encephalomyelitis (VEE) virus isolated from HeLa carrier cultures by transfection in different cell cultures have been obtained. Introduction of DNA extracted from the carrier cultures into BHK-21 cell cultures resulted in production of an actively multiplying medium-plaque virus, and parallel addition of the same DNA preparations in chick fibroblast or monkey kidney cultures led to production of small-plaque virus with a low reproduction potential. The virus produced by transfection of BHK-21 cells differed from that produced in chick fibroblast and monkey kidney cultures in electrophoretic mobility of virion envelope proteins. The infection of these cultures with virions as well as infection with genome RNA did not result in production of differing virus variants. The importance of the experimental genetic data for the problem of the nature of the infectious principle of cellular DNA preparations and of the form of existence of viral genome in chronically infected cultures with infection of the integrative type is discussed.
Asunto(s)
Virus de la Encefalitis Equina Venezolana/genética , Encefalomielitis Equina/microbiología , Variación Genética , Transfección , Animales , Embrión de Pollo , Enfermedad Crónica , Virus de la Encefalitis Equina Venezolana/aislamiento & purificación , Fibroblastos/microbiología , Haplorrinos , Células HeLa/microbiología , Humanos , Riñón , Ensayo de Placa Viral , Cultivo de VirusRESUMEN
The natural process of infection with multiploid virions of alphaviruses was modeled by inoculation of cells with artificially obtained aggregates. After infection with the aggregates the progeny of attenuated clones of Venezuelan equine encephalomyelitis virus was found to have some new properties. The virus produced larger plaques and had higher reproduction activity. In some cases, changes in the electrophoretic mobility of virion structural proteins was observed, especially a decrease in the molecular weight of the most conservative of them, C-protein. These properties were retained in passages of the virus in cell cultures. The observed phenomenon is associated with genetic interaction between defective virus genomes getting into cells as the aggregate component. Formation of multiploid virions and infection with these particles are considered as the process providing for retention of the biological properties of the virus under conditions of accumulation of DI-particles in the population.
Asunto(s)
Alphavirus/genética , Virión/genética , Animales , Embrión de Pollo , Virus de la Encefalitis Equina Venezolana/genética , Peso Molecular , Fenotipo , Ensayo de Placa Viral , Proteínas Virales/análisis , Cultivo de VirusRESUMEN
In combined paired cultivation of 8 Venezuelan equine encephalomyelitis virus, multiploid virions formed in 17.8% of cases. Five clones with this effect were used in mixed infections with clones of Semliki Forest and Sindbis viruses. In these infections changes in virions of the virus progeny were observed in 20% and 16.6%, respectively. Mixed cultivation of pairs of Sindbis and Semliki Forest viruses resulted in formation of multiploid virions in 42.8%. Besides, in two mixed populations formed upon combined multiplication of the latter viruses virions of unusual shapes were found: rounded, oval, elongated, and triangular designated as polymorphic.
Asunto(s)
Encefalomielitis Equina/microbiología , Encefalomielitis Equina Venezolana/microbiología , Infecciones por Togaviridae/microbiología , Animales , Embrión de Pollo , Células Clonales/microbiología , Virus de la Encefalitis Equina Venezolana/ultraestructura , Microscopía Electrónica , Poliploidía , Virus de los Bosques Semliki/ultraestructura , Virus Sindbis/ultraestructura , Virión/ultraestructura , Cultivo de VirusRESUMEN
Multiploid virions present in the population of multiploid-forming mutant of Venezuelan equine encephalomyelitis (VEE) virus were isolated from preparations of purified and concentrated virus by methods of zonal centrifugation in glycerin density gradient or gel filtration through biogel A-150 m. The portion of multiploid particles in such preparations reached 70%-90%. The isolated multiploid virions retained their morphology, infectivity and hemagglutinating activity. The multiploid particles were found to differ from standard virions in the sedimentation rate and buoyant density.
Asunto(s)
Virus de la Encefalitis Equina Venezolana/aislamiento & purificación , Virión/genética , Virión/aislamiento & purificación , Animales , Embrión de Pollo , Cromatografía en Gel , Virus de la Encefalitis Equina Venezolana/genética , Microscopía Electrónica , Mutación , Cultivo de VirusRESUMEN
An acute infection of HeLa cell cultures with a ts mutant of Venezuelan equine encephalomyelitis (VEE) virus may have different outcomes ranging from formation of a chronic virus-carrier state to complete elimination of virus from the culture. One of the possibilities is also a short-time carrier state (up to 10 passages) of the virus genome apparently existing in the form of DNA-provirus. The carrier state of virus genetic material in this case is indicated by the infectivity of cellular DNA and the presence in it of sequences homologous to those of virion RNA. The genetic material of VEE virus in HeLa cell cultures with signs of infection of the integrative type is also demonstrable by its complementation with exogenous ts-mutants.
Asunto(s)
Virus de la Encefalitis Equina Venezolana/patogenicidad , Mutación , Animales , Supervivencia Celular , Efecto Citopatogénico Viral , ADN/genética , Genes Virales , Células HeLa/microbiología , Humanos , Hibridación de Ácido Nucleico , ARN Viral/genética , Virulencia , Cultivo de VirusRESUMEN
Japanese encephalitis virus contained in the medium of chronically infected VERO-K43 cell cultures was shown to be sensitive to the effect of RNA-ase, resistant to treatment with urea, to have no hemagglutinating activity. After 2 passages in suckling mouse brain the virus lost its sensitivity to RNA-ase, was inactivated by urea, and its hemagglutinating activity was restored. The importance of Japanese encephalitis virus variability in the mechanism of chronic infection of cell cultures is discussed.