RESUMEN
The sequences of the genomes in the TO-336 vaccine strain (TO-336vac) of rubella virus and its wild progenitor virus (TO-336wt) have been determined and compared with each other. There were 21 differences in the nucleotide sequences between the TO-336vac and the TO-336wt: 13 in the nonstructural protein open reading frame (NSP-ORF), five in the structural protein open reading frame (SP-ORF) and three in the untranslated regions (UTRs) (one in each three UTRs). These mutations resulted in amino acid substitutions at ten residues. Of the ten substitutions, eight were in NSP-ORF and two were in the SP-ORF. Of the eight substitutions in NSP-ORF, four (amino acids (aa) 320, 501, 573 and 704) were in the regions of unknown function, two (aa 1154 and 1159) were within the protease motif, and two (aa 1351 and 1559) were within the helicase motif. Both of the two residues (aa 890 and 954) in the SP-ORF were within the E1 gene. The predicted second structure of the 5'UTR of the TO-336vac was identical to that of TO-336wt. Comparing the TO-336 sequences with other four strains, Therien and M33 (wild viruses), and RA27/3 and Cendehill (vaccine viruses), the mutations responsible for attenuation are thought to differ with each vaccine strain. This is the first report of sequencing in a pair of live attenuated rubella vaccines and their wild-type parent.
Asunto(s)
Mutación , Provirus/genética , Vacuna contra la Rubéola , Virus de la Rubéola/genética , Secuencia de Bases , Datos de Secuencia Molecular , Sistemas de Lectura Abierta , Filogenia , Vacunas AtenuadasRESUMEN
UNLABELLED: A boy with congenital rubella syndrome developed dysgammaglobulinaemia with elevated serum levels of IgM. CD154 was not induced on his peripheral blood mononuclear cells when rubella virus RNA was detected in his throat swabs and peripheral blood by reverse transcriptase polymerase chain reaction. Following intravenous immunoglobulin therapy, improvement of immunoglobulin abnormalities, disappearance of rubella virus and normalisation of CD154 expression were demonstrated. CONCLUSION: These findings implicate the efficacy of intravenous immunoglobulin therapy for dysgammaglobulinaemia in congenital rubella syndrome and a role of CD154 for a prolonged virus infection.
Asunto(s)
Anticuerpos Antivirales/sangre , Ligando de CD40/inmunología , Disgammaglobulinemia/inmunología , Inmunoglobulinas Intravenosas/uso terapéutico , Síndrome de Rubéola Congénita/tratamiento farmacológico , Virus de la Rubéola/aislamiento & purificación , Disgammaglobulinemia/etiología , Regulación de la Expresión Génica , Humanos , Inmunoglobulina M , Inmunoglobulinas Intravenosas/administración & dosificación , Lactante , Masculino , ARN Viral/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Síndrome de Rubéola Congénita/complicaciones , Síndrome de Rubéola Congénita/inmunología , Virus de la Rubéola/genética , Virus de la Rubéola/inmunología , Resultado del TratamientoRESUMEN
The repeating sequences of the toxin A gene from toxin A-negative, toxin B-positive (toxin A-, toxin B+) strains of Clostridium difficile which were isolated in geographically separated facilities in Japan and Indonesia were determined. All six strains tested had identical repeating sequences with two deletions (1548 and 273 nucleotides in size) in the toxin A gene. A PCR method was designed to detect the deletions and the deletions were confirmed in all 50 toxin A-, toxin B+ strains examined by this method. Western immunoblot analysis revealed that polyclonal antiserum against native toxin A did not react with the concentrated culture filtrates of the toxin A-, toxin B+ strains. These results may suggest that toxin A-, toxin B+ strains have deletions of the two thirds of the repeating regions of the toxin A gene, which encodes the epitopes fully responsible for the reaction with the polyclonal antiserum.
Asunto(s)
Proteínas Bacterianas , Toxinas Bacterianas/metabolismo , Clostridioides difficile/genética , Enterocolitis Seudomembranosa/microbiología , Enterotoxinas/genética , Eliminación de Secuencia , Adulto , Secuencia de Aminoácidos , Niño , Clostridioides difficile/metabolismo , Electroforesis en Gel de Poliacrilamida , Genes Bacterianos , Humanos , Immunoblotting , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Análisis de Secuencia de ADN , Transcripción GenéticaAsunto(s)
Linfocitos T CD8-positivos/inmunología , Virus de la Rubéola/aislamiento & purificación , Rubéola (Sarampión Alemán)/inmunología , Líquido Sinovial/virología , Adulto , ADN Viral/análisis , Femenino , Genoma Viral , Humanos , Virus de la Rubéola/genética , Líquido Sinovial/inmunología , Subgrupos de Linfocitos T/inmunologíaRESUMEN
E1 gene nucleotide sequences of 63 rubella virus isolates from North America, Europe, and Asia isolated between 1961 and 1997 were compared phylogenetically. Two genotypes were evident: Genotype I contained 60 viruses from North America, Europe, and Japan, and genotype II contained 3 viruses from China and India. The genotype I isolates prior to 1970 grouped into a single diffuse clade, indicating intercontinental circulation, while most post-1975 viruses segregated into geographic clades from each continent, indicating evolution in response to vaccination programs. The E1 amino acid sequences differed by no more than 3%; thus, no major antigenic variation was apparent. Among 4 viruses from congenital rubella syndrome that occurred following reinfection, only one amino acid substitution occurred in several important epitopes, indicating that antigenic drift is not important in this phenomenon. However, 2 viruses isolated from chronic arthritis exhibited changes in these epitopes. Isolates of the RA 27/3 vaccine strain were readily identifiable by nucleotide sequence.
Asunto(s)
Virus de la Rubéola/genética , Rubéola (Sarampión Alemán)/epidemiología , Rubéola (Sarampión Alemán)/virología , Secuencia de Aminoácidos , Asia/epidemiología , Europa (Continente)/epidemiología , Humanos , Datos de Secuencia Molecular , América del Norte/epidemiología , Filogenia , Virus de la Rubéola/clasificación , Virus de la Rubéola/aislamiento & purificación , Proteínas del Envoltorio Viral/genéticaRESUMEN
Fetal infection with rubella virus was diagnosed based on the detection of viral genome in the fetus-derived tissues. While viral genomes were detected in 41 of those 112 cases (36.7%) where rubella virus infection of the mother was apparent, only 7 of 141 cases (5. 0%) showed evidence of fetal infection when maternal rubella infection was inapparent. All 184 babies born of genome-negative mothers have no congenital disorder, while 2 out of 7 genome-positive babies have a congenital disorder (28.6%). Rubella virus was not transmitted across the placenta when infection occurred prior to gestation. Transmission rate increased to a maximum level during the first trimester and declined to 0% until 20 weeks of gestation. Interval of viral transmission from the onset of rash in the mother was about 10 days to the placental villi and 20-30 days to the fetus. A phylogenetic tree of 61 virus isolates suggested no difference of virulence/teratogenicity among the virus isolates.
Asunto(s)
Enfermedades Fetales/diagnóstico , Complicaciones Infecciosas del Embarazo/diagnóstico , Diagnóstico Prenatal , Síndrome de Rubéola Congénita/diagnóstico , Virus de la Rubéola/genética , Rubéola (Sarampión Alemán)/diagnóstico , Femenino , Enfermedades Fetales/virología , Humanos , Recién Nacido , Transmisión Vertical de Enfermedad Infecciosa , Japón/epidemiología , Epidemiología Molecular , Filogenia , Reacción en Cadena de la Polimerasa/métodos , Embarazo , Complicaciones Infecciosas del Embarazo/virología , Resultado del Embarazo , ARN Viral/aislamiento & purificación , Rubéola (Sarampión Alemán)/epidemiología , Rubéola (Sarampión Alemán)/virología , Virus de la Rubéola/aislamiento & purificaciónRESUMEN
Strain-specific nucleotide sequences of E1 and NS4 genes in five strains of a live rubella virus vaccine manufactured in Japan were identified for comparison, using 2389 nucleotides (1443 nucleotides of the E1 gene, 41 of the 3' terminal region following the E1 gene and 905 of the NS4 gene). Sequences of the E1 gene in three strains (Matsuura, TCRB19 and To-336) were identified. Takahashi and Matsuba strains shared common sequences, but were discriminated by the sequence of the NS4 gene. These five strains showed a phylogenetic relationship with the places of their isolation. In a comparative study of three strains with their unattenuated progenitors, the nucleotides in these regions were almost conserved during the attenuation process.
Asunto(s)
Genes Virales , Vacuna contra la Rubéola/genética , Virus de la Rubéola/genética , Proteínas del Envoltorio Viral/genética , Proteínas no Estructurales Virales/genética , Secuencia de Bases , Japón , Datos de Secuencia Molecular , Virus de la Rubéola/clasificación , Especificidad de la EspecieRESUMEN
Twenty-six strains of rubella virus were compared with each other for a molecular epidemiologic study of the virus in three East Asian countries, using the E1 gene of 1443 nucleotides and the following 41 nucleotides in a noncoding region. Nucleotide substitution rates among strains were 0.0-9.4/100 nucleotides. A phylogenetic tree drawn indicated that 2 of 3 Chinese strains were quite different from the other 24 strains; all isolates in the 1960s were classified into a single group independent of the place of isolation, which includes isolates from Japan, the United States, and the United Kingdom; 11 strains of Japanese isolates collected during 1976-1991 made one subbranch derived from the 1960s group; and 2 isolates from the northeast part of Japan in 1990 made a third but minor unique branch. Therefore, at least two groups of the virus cocirculated in Japan around 1990. Antigenic variation of the virus was very small among these strains.
Asunto(s)
Antígenos Virales/genética , Virus de la Rubéola/genética , Rubéola (Sarampión Alemán)/virología , Proteínas del Envoltorio Viral/genética , Secuencia de Aminoácidos , Secuencia de Bases , China/epidemiología , ADN Viral , Asia Oriental/epidemiología , Genes Virales , Humanos , Japón/epidemiología , Epidemiología Molecular , Datos de Secuencia Molecular , Filogenia , Rubéola (Sarampión Alemán)/epidemiología , Virus de la Rubéola/clasificación , Homología de Secuencia de Ácido NucleicoRESUMEN
OBJECTIVE: Our purpose was to develop a reliable method for prenatal diagnosis of fetal rubella infection through the detection of viral ribonucleic acid extracted from the chorionic villi, amniotic fluid, or fetal blood in pregnant women. STUDY DESIGN: Double amplification of rubella viral ribonucleic acid by nested polymerase chain reaction after reverse transcription was applied to samples from 34 women suspected of having rubella. The results were compared with those of serum antibody and levels of rubella virus-specific immunoglobulin M antibodies in fetal blood. RESULTS: Viral ribonucleic acid was revealed in 8 of 34 cases (23.5%). In the remaining 26 cases, healthy babies were born in 24, 1 was electively aborted, and 1 died in the thirty-sixth week of pregnancy of unknown causes. CONCLUSIONS: This method allowed very early detection of fetal rubella infection by sampling of chorionic villi and amniotic fluid compared with evaluation of the maternal symptoms and serum antibody levels. Fetal blood was also more useful for making a diagnosis up to the twentieth week of pregnancy than was measuring rubella virus-specific immunoglobulin M antibodies.
Asunto(s)
Enfermedades Fetales/diagnóstico , Reacción en Cadena de la Polimerasa , Diagnóstico Prenatal , ARN Viral/sangre , Virus de la Rubéola/genética , Rubéola (Sarampión Alemán)/diagnóstico , Adulto , Anticuerpos Antivirales/sangre , Femenino , Sangre Fetal/virología , Humanos , Inmunoglobulina M/sangre , Embarazo , Resultado del Embarazo , ADN Polimerasa Dirigida por ARN , Rubéola (Sarampión Alemán)/embriología , Virus de la Rubéola/inmunologíaRESUMEN
To assess the potential of an infectious Sindbis virus vector (dsSIN) as a eukaryotic expression vector, dsSIN recombinants which contain each of the rubella virus (RUB) structural proteins (C, E2, and E1) individually were constructed. Expression of the RUB proteins by each dsSIN recombinant was robust and processing was similar to that observed when these proteins were expressed using other vectors. The C and E2 recombinants, each of which contains a cassette of less than 1,000 nts, were stable through low multiplicity amplification; however, the E1 recombinant, which contains a 1700 nt cassette, was not. Therefore, use of the E1 recombinant is restricted to stock derived from the original transfection. The replication rate of dsSIN and the C and E2 recombinants was similar, however, the replication rate of the E1 recombinant was slower. No phenotypic mixing of any of the RUB proteins in recombinant dsSIN virions could be detected.
Asunto(s)
Antígenos Virales/genética , Virus de la Rubéola/genética , Virus Sindbis/genética , Proteínas Estructurales Virales/genética , Animales , Secuencia de Bases , Western Blotting , Células Cultivadas , Chlorocebus aethiops , Cartilla de ADN/química , Vectores Genéticos , Glicoproteínas/biosíntesis , Datos de Secuencia Molecular , ARN Viral/biosíntesis , Proteínas Recombinantes , Factores de Tiempo , Células Vero , Proteínas Estructurales Virales/inmunologíaRESUMEN
BACKGROUND: We examined an infant with congenital rubella syndrome (CRS). The purpose of this report is to describe the skin manifestations in this patient and to prove that they were associated with rubella virus. OBSERVATIONS: A 7-month-old boy presented with generalized brownish macules, indurated erythemas, papules, and pigmentation. They first appeared at around 3 months of age. His mother had contracted rubella during the 14th gestational week. At the time of examination, rubella-specific IgM antibody was positive in both serum and cerebrospinal fluid of the baby. A physical exam had revealed deafness, mental and physical retardation, interstitial pneumonitis, and hepatosplenomegaly. A skin biopsy specimen showed a dense infiltration mainly of lymphocytes, with B cells predominant in the deep dermis. Electron microscopically abundant tubuloreticular structures were observed in capillary endothelial cells, lymphocytes, and dermal fibroblasts. Polymerase chain reaction (PCR) analysis suggested that rubella virus RNA was present in the patient's skin specimen, cerebrospinal fluid, and total blood. CONCLUSIONS: The cutaneous manifestations of our patient were extraordinary and informative. These prominent skin lesions should be recognized as cutaneous markers of CRS.
Asunto(s)
Síndrome de Rubéola Congénita/patología , Virus de la Rubéola/aislamiento & purificación , Pigmentación de la Piel , Piel/patología , Humanos , Lactante , Masculino , Reacción en Cadena de la Polimerasa , ARN Viral/análisis , Síndrome de Rubéola Congénita/microbiología , Piel/microbiologíaRESUMEN
Formation, accumulation, and storage of two components of the Oryzias latipes hatching enzyme, high and low choriolytic enzymes (HCE and LCE), were examined by immunocytochemical and immunoblotting methods. Both of the enzymes were found to be formed specifically in the hatching gland cells at the stages of lens formation to eye pigmentation and their accumulation proceeded markedly and concurrently up to Day 5.5 embryos (the stage just before hatching). The amount of HCE formed was more abundant than that of LCE. In the hatching gland cells, HCE and LCE were found to be packaged in the same secretory granules but in distinct arrangement; HCE is localized to the inside of granules whereas LCE is situated at the periphery of the same granules. Their segregated arrangement is compatible with their relative quantities formed per embryo. The results provide not only the cellular and developmental basis for a view that this hatching enzyme is an enzyme system composed of HCE and LCE but also a clue to the regulatory mechanism of concurrent syntheses of two different specific proteins in the same embryonic cell.
Asunto(s)
Gránulos Citoplasmáticos/enzimología , Embrión no Mamífero/enzimología , Endopeptidasas/metabolismo , Metaloendopeptidasas/metabolismo , Oryzias/embriología , Animales , Especificidad de Anticuerpos , Western Blotting , Endopeptidasas/biosíntesis , Inmunohistoquímica , Metaloendopeptidasas/biosíntesis , Metaloendopeptidasas/inmunologíaRESUMEN
High choriolytic enzyme (HCE), a constituent protease of the hatching enzyme of the teleost, Oryzias latipes, swells its natural substrate, egg envelope (chorion) by hydrolyzing it partially. This enzyme was found to be bound tightly to the chorion when it exerted catalytic action. This was evidenced by the experimental results showing (i) that the turnover of this enzyme seemed to be hindered by the chorion, (ii) that the enzyme bound to the chorion could be recovered by washing with an alkaline medium, and (iii) that the bound enzyme could be quantified by radioimmunological estimation. The bound enzyme sustained its original activity and the binding between the enzyme and the chorion seems to be stoichiometric.
Asunto(s)
Corion/metabolismo , Proteínas del Huevo/metabolismo , Metaloendopeptidasas/metabolismo , Animales , Sitios de Unión , Catálisis , Hidrólisis , Cinética , Ratones , Ratones Endogámicos BALB C , Oryzias , Radioinmunoensayo , Especificidad por SustratoRESUMEN
Single-serum diagnosis of recent rubella infection was attempted with the use of hemagglutination inhibition (HI) test and two commercially available enzyme-linked immunosorbent assays (ELISAs). The period during which IgM antibody was detectable by IgM capture ELISA was 4 to 30 days after the onset of rash. Rubella IgG ELISA values relative to HI titer were lower in the sera from the patients with recent infection than in the sera from the subjects with remote infection. IgM ELISA and the combined use of IgG ELISA and HI test are two useful methods of single-serum diagnosis of recent rubella infection.
Asunto(s)
Ensayo de Inmunoadsorción Enzimática , Pruebas de Inhibición de Hemaglutinación , Rubéola (Sarampión Alemán)/diagnóstico , Anticuerpos Antivirales/análisis , Afinidad de Anticuerpos , Estudios de Evaluación como Asunto , Humanos , Inmunoglobulina G/análisis , Inmunoglobulina M/análisis , Rubéola (Sarampión Alemán)/inmunología , Factores de TiempoRESUMEN
Fusion of rubella virus-infected cells was induced by their brief treatment at pH below 6.0. Exposure of rubella virus to pH 5 caused an irreversible conformational change of the viral envelope glycoproteins, E1 and E2. The change was manifested in the marked reduction in both infectivity and haemagglutinating activity of the virus, the increased resistance of E1 and decreased resistance of E2 polypeptides to proteolytic digestion with trypsin, and the acquisition of liposome-binding activity of the virus. The above changes are presumed to mimic the events occurring in the acidic environment within endosomes following endocytosis of the virus.
Asunto(s)
Virus de la Rubéola/ultraestructura , Proteínas del Envoltorio Viral/ultraestructura , Hemaglutininas Virales/fisiología , Concentración de Iones de Hidrógeno , Liposomas , Peso Molecular , Pruebas de Precipitina , Conformación Proteica , Tripsina/metabolismo , Proteínas Virales de Fusión/ultraestructuraRESUMEN
Hemagglutinating (HA) activity of rubella virus was inactivated with 2-mercaptoethanol (2ME) in a dose-dependent manner. But even low concentrations of 2ME, which had little effect on HA activity by themselves, greatly increased the sensitivity of spike polypeptides to the subsequent trypsin treatment. Increased trypsin sensitivity was shown by an enhanced reduction of HA activity and an enhanced proteolytic removal of both E1 and E2 polypeptides from the surface of the virion. The findings indicate that 2ME causes an extensive disruption in the conformation of spikes composed of E1 and E2 polypeptides.