RESUMEN
We previously described the use of an established reverse genetics system for the generation of recombinant human influenza A viruses from cloned cDNAs. Here, we have assembled a set of plasmids to allow recovery of the avian H5N1 influenza virus A/Turkey/England/50-92/91 entirely from cDNA. This system enables us to introduce mutations or truncations into the cDNAs to create mutant viruses altered specifically in a chosen gene. These mutant viruses can then be used in future pathogenesis studies in chickens and in studies to understand the host range restrictions of avian influenza viruses in humans.
Asunto(s)
Técnicas Genéticas , Subtipo H5N1 del Virus de la Influenza A/genética , Animales , Línea Celular , Cricetinae , ADN Complementario , ADN Viral , ARN Polimerasas Dirigidas por ADN/metabolismo , Perros , HumanosRESUMEN
Previously we described a heterosexual outbreak of HIV-1 subtype B in a town in the north of England (Doncaster) where 11 of 13 infections were shown to be linked by phylogenetic analysis of the env gp120 region. The 11 infections were related to a putative index case, Don1, and further divided into two groups based on the patients' disease status, their viral sequences, and other epidemiological information. Here we describe two further findings. First, we found that viral isolates and gp120 recombinant viruses derived from patients from one group used the CCR5 coreceptor, whereas viruses from the other group could use both the CCR5 and CXCR4 coreceptors. Patients with the X4/R5 dual tropic strains were symptomatic when diagnosed and progressed rapidly, in contrast to the other patient group that has remained asymptomatic, implying a link between the tropism of the strains and disease outcome. Second, we present additional sequence data derived from the index case, demonstrating the presence of sequences from both clades, with an average interclade distance of 9.56%, providing direct evidence of a genetic link between these two groups. This new study shows that Don1 harbored both strains, implying he was either dually infected or that over time intrahost diversification from the R5 to R5/X4 phenotype occurred. These events may account for/have led to the spread of two genetically related strains with different pathogenic properties within the same heterosexual community.
Asunto(s)
Infecciones por VIH/fisiopatología , Infecciones por VIH/virología , VIH-1/genética , VIH-1/patogenicidad , Receptores del VIH/fisiología , Secuencia de Aminoácidos , Progresión de la Enfermedad , Inglaterra , Infecciones por VIH/epidemiología , VIH-1/clasificación , VIH-1/aislamiento & purificación , Heterosexualidad , Humanos , Datos de Secuencia Molecular , Filogenia , Receptores CCR5/metabolismo , Receptores CXCR4/metabolismo , Receptores CXCR4/fisiología , Homología de SecuenciaRESUMEN
BACKGROUND: Cytotoxic T lymphocytes have been shown to reduce viraemia during acute HIV-1 infection; however the role of neutralizing antibodies in this process is unclear. One confounding factor may be artefacts introduced by viral culture. OBJECTIVE: To assess the development of autologous neutralizing and non-neutralizing antibodies following acute HIV-1 infection using recombinant viruses with envelopes amplified directly from patient peripheral blood mononuclear cells, thereby avoiding in vitro selection. METHODS: Disease progression in four homosexual men was monitored from acute infection for up to 2.5 years, in the absence of antiretroviral therapy. Antibodies to viral envelope protein were quantified by enzyme-linked immunosorbent assay. Development of neutralizing antibodies was monitored using a quantitative infectivity reduction assay, sequential serum, recombinant viruses and target cells with defined receptor expression. RESULTS: The time to development of neutralizing antibodies after onset of symptoms was 3, 5, 7 and 16 months in the four patients. There was no correlation between development of neutralizing antibodies and the resolution of viraemia in any of the patients. However, antibodies to the envelope were detectable as early as 2 weeks after onset of symptoms. CONCLUSIONS: Neutralizing antibodies do not contribute to the control of viraemia in acute HIV-1 infection. However, antibodies to the envelope could be detected at the time of reduction in plasma viraemia and so other effector functions of antibodies may play a role in viral clearance.