RESUMEN
Dengue virus is the most significant virus transmitted by arthropods worldwide and may cause a potentially fatal systemic disease named dengue hemorrhagic fever. In this work, dengue virus serotype 4 was detected in the tissues of one fatal dengue hemorrhagic fever case using electron immunomicroscopy and molecular methods. This is the first report of dengue virus polypeptides findings by electron immunomicroscopy in human samples. In addition, not-previously-documented virus-like particles visualized in spleen, hepatic, brain, and pulmonary tissues from a dengue case are discussed.
Asunto(s)
Virus del Dengue/genética , Virus del Dengue/ultraestructura , Dengue Grave/diagnóstico , Adulto , Anticuerpos Antivirales/sangre , Encéfalo/ultraestructura , Encéfalo/virología , Cuba , ADN Viral/análisis , Virus del Dengue/aislamiento & purificación , Ensayo de Inmunoadsorción Enzimática , Resultado Fatal , Femenino , Corazón/virología , Humanos , Inmunoglobulina M/sangre , Riñón/ultraestructura , Riñón/virología , Hígado/ultraestructura , Hígado/virología , Microscopía Electrónica de Transmisión/métodos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Dengue Grave/virología , Bazo/ultraestructura , Bazo/virologíaRESUMEN
Acute and late convalescent sera (collected at day 5 of disease onset and 1 year later) from dengue fever (DF) and dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS) laboratory confirmed cases, were tested for antibody-dependent cell-mediated cytotoxicity (ADCC) activity using dengue 1 (DENV-1) or dengue 2 (DENV-2) infected cells as target. All patients experienced their first dengue virus (DENV) infection 20 years before. ADCC activity was detected in acute sera from DHF/DSS but not in sera from DF patients. However, 1 year after illness, ADCC activity was observed in all cases. This preliminary report represents one of the few studies of ADCC in dengue patients and suggests that ADCC could be implicated in dengue pathogenesis.
Asunto(s)
Anticuerpos Antivirales/inmunología , Citotoxicidad Celular Dependiente de Anticuerpos , Virus del Dengue/inmunología , Dengue/inmunología , Línea Celular , Células Cultivadas , Pruebas Inmunológicas de Citotoxicidad , Humanos , Leucocitos Mononucleares/inmunologíaRESUMEN
It was recently reported that disease severity increased during the 1997 Cuban dengue 2 virus epidemic and it was suggested that this might be explained by the appearance of neutralization resistant escape mutants. We investigated these observations and ideas by sequencing 20 dengue 2 virus isolates obtained during the early (low case fatality rate) and the late (high case fatality rate) phases of the outbreak. Our results showed total conservation of the E gene sequence for these isolates suggesting that the selection of envelope gene escape mutants was not the determinant of increased disease severity. Alignment of these sequences with those available in GenBank, followed by Maximum likelihood phylogenetic analysis generated a tree, which indicated that our isolates are closely related to the virus that circulated in Venezuela in 1997/98 and subsequently in Martinique in 1998. This "American/Asian" genotype has therefore gradually dispersed across the Caribbean region during the past 5 years.
Asunto(s)
Virus del Dengue/genética , Dengue/epidemiología , Brotes de Enfermedades , Proteínas del Envoltorio Viral/genética , Cuba/epidemiología , Dengue/mortalidad , Virus del Dengue/aislamiento & purificación , Humanos , Epidemiología Molecular , Datos de Secuencia Molecular , Filogenia , Proteínas del Envoltorio Viral/análisisRESUMEN
We have expressed a recombinant Dengue 4 virus envelope glycoprotein (E4rec), truncated at its C-terminus by 53 amino acids, in Pichia pastoris. The presence of E4rec was confirmed by Western-blot using anti-DEN 4 hyper immune mouse ascitic fluid. E4rec migrated during SDS-PAGE as a 64 kDa protein. Treatment with endoglycosidases showed that the E protein was modified by the addition of short mannose chains and the absence of hyperglycosylation. When administered to BALB-C mice, E4rec elicited a DEN 4 neutralizing antibody response haemagglutination inhibition antibodies and specific memory T cell response. Mice immunized were also significantly protected against lethal DEN 4 virus challenge (86.6%, p < 0.001).
Asunto(s)
Anticuerpos Antivirales/sangre , Virus del Dengue/inmunología , Vacunas Sintéticas/inmunología , Proteínas del Envoltorio Viral/inmunología , Vacunas Virales/inmunología , Animales , Dengue/prevención & control , Inmunización , Ratones , Ratones Endogámicos BALB C , Pruebas de Neutralización , Pichia/genética , Proteínas Recombinantes/inmunologíaRESUMEN
A rapid centrifugation assay was compared with conventional tube cell culture for dengue virus isolation in both sera and autopsy samples from dengue and dengue hemorrhagic fever/dengue shock syndrome fatal cases. The rapid centrifugation assay allowed isolation of virus from 16.6% more samples than the conventional method, and it shortened the time for dengue virus detection. Finally, it allowed the isolation of dengue 2 virus in 42.8% of tissue samples from five fatal cases. Our results suggest that the rapid centrifugation assay may be useful for detection of dengue virus in clinical specimens.
Asunto(s)
Centrifugación , Virus del Dengue/crecimiento & desarrollo , Virus del Dengue/aislamiento & purificación , Dengue/diagnóstico , Dengue Grave/diagnóstico , Virología/métodos , Aedes , Animales , Células Cultivadas , Dengue/sangre , Dengue/virología , Humanos , Dengue Grave/sangre , Dengue Grave/virología , Cultivo de VirusRESUMEN
Papers dealing with the study of the immune response to dengue virus infection and with the role it may play in the pathogenesis of dengue hemorrhaghic fever are increasingly important. The memory human T cell response in individuals with history of dengue infection during the Cuban epidemic was studied in this paper for a further evaluation of the antigenicity of viral proteins. To this end, mononuclear cells of peripheral blood from individuals immune to dengue and from a group of control subjects with viral antigens were incubated. It was obtained a significant proliferative response of lymphocytes from individuals with history of infection against dengue virus type 2 compared with control subjects. It was proved this way the dengue virus-specific memory T CD4+ cell response in the individuals under study.
Asunto(s)
Anticuerpos Antivirales/sangre , Antígenos Virales/inmunología , Virus del Dengue/inmunología , Dengue/sangre , Dengue/inmunología , Linfocitos T/inmunología , Adulto , Antígenos Virales/sangre , Femenino , Humanos , Masculino , Persona de Mediana EdadRESUMEN
4 primer sets, were used to allow the amplification of a nucleotide sequence with unique size for each of the dengue virus serotypes by polymerase chain reaction (PRC). The method consisted in the extraction of ribonucleic acid from supernatant of infected cell cultures, reverse transcription-polymerase chain reaction (RT-PCR). This was completed in approximately 7 hours in a simple tube. The size of the amplified sequence was evidenced by electrophoresis in Agarose gel stained with ethidium. The method showed a sensitivity of at least 2.5 plate forming units (pfu) per tube of reaction. It es useful for the detection and simultaneous identification of the 4 serotypes, starting from supernatants of infected strains cultures from different countries of the Caribbean, Central America, and South America.