Caracterización del virus de la fiebre aftosa atenuado para bovinos por pasajes en embrión de pollo

La obtención natural o artificial de mutantes es una propiedad común a todos los virus. Estas mutantes se pueden caracterizar por su sensibilidad a la tenperatura, por su patogenicidad en diferentes animales, por su especificidad hacia un determinado huésped, etc.. Cepas atenuadas aisladas en distintos sistemas virales han sido utilizadas como agentes vacunantes con resultados muy alentadores. Actualmente, se están empleando en Venezuela cepas de virus de la fiebre aftosa atenuadas para bovinos en la producción de vacunas. Estas cepas han sido utilizadas teniendo en cuenta su bajo grado de patogenicidad y alta calidad inmunogénica. Sin embargo, el criterio por el cual fueron seleccionadas es altamente empírico ya que se conoce muy poco acerca del mecanismo de atenuación. Principalmente, habian sido estudiadas en cuanto a sus caracteristicas biológicas de multiplicación respecto de las cepas virulentas. En base a lo previamente expuesto, se plantearon los siguientes objetivos para este tema de tesis: 1°) El estudio de los factores celulares y/o virales que influyen en el mecanismo de atenuación. 2°) Las caracteristicas inhibitorias de la multiplicación del virus salvaje por las cepas atenuadas. Estos objetivos fueron enfocados a traves de tres puntos principales: - Caracterización biológica del virus atenuado (infecciosidad, ciclo de multiplicación, etc.). - Caracterización de la interferencia por el virus atenuado en la multiplicación del virus salvaje. - Caracterización fisiquuímica del virus atenuado y de sus componentes (coeficiente de sedimentación, densidad, análisis de su genoma).Fil: Polacino, Patricia Susana. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina

Pathogenic infection of Macaca nemestrina with a CCR5-tropic subtype-C simian-human immunodeficiency virus

Background: Although pig-tailed macaques (Macaca nemestrina) have been used in AIDS research for years, less is known about the early immunopathogenic events in this species, as compared to rhesus macaques (Macaca mulatta). Similarly, the events in early infection are well-characterized for simian immunodeficiency viruses (SIV), but less so for chimeric simian-human immunodeficiency viruses (SHIV), although the latter have been widely used in HIV vaccine studies. Here, we report the consequences of intrarectal infection with a CCR5-tropic clade C SHIV-1157ipd3N4 in pig-tailed macaques. Results: Plasma and cell-associated virus was detectable in peripheral blood and intestinal tissues of all four pig-tailed macaques following intrarectal inoculation with SHIV-1157ipd3N4. We also observed a rapid and irreversible loss of CD4+ T cells at multiple mucosal sites, resulting in a marked decrease of CD4:CD8 T cell ratios 0.5–4 weeks after inoculation. This depletion targeted subsets of CD4+ T cells expressing the CCR5 coreceptor and having a CD28-CD95+ effector memory phenotype, consistent with the R5-tropism of SHIV-1157ipd3N4. All three animals that were studied beyond the acute phase seroconverted as early as week 4, with two developing cross-clade neutralizing antibody responses by week 24. These two animals also demonstrated persistent plasma viremia for >48 weeks. One of these animals developed AIDS, as shown by peripheral blood CD4+ T-cell depletion starting at 20 weeks post inoculation. Conclusion: These findings indicate that SHIV-1157ipd3N4-induced pathogenesis in pig-tailed macaques followed a similar course as SIV-infected rhesus macaques. Thus, R5 SHIV-C-infection of pig-tailed macaques could provide a useful and relevant model for AIDS vaccine and pathogenesis research

Dramatic Rise in Plasma Viremia after CD8+ T Cell Depletion in Simian Immunodeficiency Virus–infected Macaques

To determine the role of CD8+ T cells in controlling simian immunodeficiency virus (SIV) replication in vivo, we examined the effect of depleting this cell population using an anti-CD8 monoclonal antibody, OKT8F. There was on average a 99.9% reduction of CD8 cells in peripheral blood in six infected Macaca mulatta treated with OKT8F. The apparent CD8 depletion started 1 h after antibody administration, and low CD8 levels were maintained until day 8. An increase in plasma viremia of one to three orders of magnitude was observed in five of the six macaques. The injection of a control antibody to an infected macaque did not induce a sustained viral load increase, nor did it significantly reduce the number of CD8+ T cells. These results demonstrate that CD8 cells play a crucial role in suppressing SIV replication in vivo

TRIM5α Modulates Immunodeficiency Virus Control in Rhesus Monkeys

The cytoplasmic TRIM5α proteins of certain mammalian lineages efficiently recognize the incoming capsids of particular retroviruses and potently restrict infection in a species-specific manner. Successful retroviruses have evolved capsids that are less efficiently recognized by the TRIM5α proteins of the natural hosts. To address whether TRIM5α contributes to the outcome of retroviral infection in a susceptible host species, we investigated the impact of TRIM5 polymorphisms in rhesus monkeys on the course of a simian immunodeficiency virus (SIV) infection. Full-length TRIM5α cDNAs were derived from each of 79 outbred monkeys and sequenced. Associations were explored between the expression of particular TRIM5 alleles and both the permissiveness of cells to SIV infection in vitro and clinical sequelae of SIV infection in vivo. Natural variation in the TRIM5α B30.2(SPRY) domain influenced the efficiency of SIVmac capsid binding and the in vitro susceptibility of cells from the monkeys to SIVmac infection. We also show the importance in vivo of the interaction of SIVmac with different allelic forms of TRIM5, demonstrating that particular alleles are associated with as much as 1.3 median log difference in set-point viral loads in SIVmac-infected rhesus monkeys. Moreover, these allelic forms of TRIM5 were associated with the extent of loss of central memory (CM) CD4+ T cells and the rate of progression to AIDS in the infected monkeys. These findings demonstrate a central role for TRIM5α in limiting the replication of an immunodeficiency virus infection in a primate host

Lack of viral control and development of combination antiretroviral therapy escape mutations in macaques after bone marrow transplantation

We have previously demonstrated robust control of simian/human immunodeficiency virus (SHIV1157-ipd3N4) viremia following administration of combination antiretroviral therapy (cART) in pigtailed macaques. Here, we sought to determine the safety of hematopoietic stem cell transplantation (HSCT) in cART-suppressed and unsuppressed animals

Vaccination against Heterologous R5 Clade C SHIV: Prevention of Infection and Correlates of Protection

A safe, efficacious vaccine is required to stop the AIDS pandemic. Disappointing results from the STEP trial implied a need to include humoral anti-HIV-1 responses, a notion supported by RV144 trial data even though correlates of protection are unknown. We vaccinated rhesus macaques with recombinant simian immunodeficiency virus (SIV) Gag-Pol particles, HIV-1 Tat and trimeric clade C (HIV-C) gp160, which induced cross-neutralizing antibodies (nAbs) and robust cellular immune responses. After five low-dose mucosal challenges with a simian-human immunodeficiency virus (SHIV) that encoded a heterologous R5 HIV-C envelope (22.1% divergence from the gp160 immunogen), 94% of controls became viremic, whereas one third of vaccinees remained virus-free. Upon high-dose SHIV rechallenge, all controls became infected, whereas some vaccinees remained aviremic. Peak viremia was inversely correlated with both cellular immunity (p<0.001) and cross-nAb titers (p<0.001). These data simultaneously linked cellular as well as humoral immune responses with the degree of protection for the first time

Dynamics of Envelope Evolution in Clade C SHIV-Infected Pig-Tailed Macaques during Disease Progression Analyzed by Ultra-Deep Pyrosequencing

Understanding the evolution of the human immunodeficiency virus type 1 (HIV-1) envelope during disease progression can provide tremendous insights for vaccine development, and simian-human immunodeficiency virus (SHIV) infection of non-human primate provides an ideal platform for such studies. A newly developed clade C SHIV, SHIV-1157ipd3N4, which was able to infect rhesus macaques, closely resembled primary HIV-1 in transmission and pathogenesis, was used to infect several pig-tailed macaques. One of the infected animals subsequently progressed to AIDS, whereas one remained a non-progressor. The viral envelope evolution in the infected animals during disease progression was analyzed by a bioinformatics approach using ultra-deep pyrosequencing. Our results showed substantial envelope variations emerging in the progressor animal after the onset of AIDS. These envelope variations impacted the length of the variable loops and charges of different envelope regions. Additionally, multiple mutations were located at the CD4 and CCR5 binding sites, potentially affecting receptor binding affinity, viral fitness and they might be selected at late stages of disease. More importantly, these envelope mutations are not random since they had repeatedly been observed in a rhesus macaque and a human infant infected by either SHIV or HIV-1, respectively, carrying the parental envelope of the infectious molecular clone SHIV-1157ipd3N4. Moreover, similar mutations were also observed from other studies on different clades of envelopes regardless of the host species. These recurring mutations in different envelopes suggest that there may be a common evolutionary pattern and selection pathway for the HIV-1 envelope during disease progression

Caracterización del virus de la fiebre aftosa atenuado para bovinos por pasajes en embrión de pollo

La obtención natural o artificial de mutantes es una propiedad común a todos los virus. Estas mutantes se pueden caracterizar por su sensibilidad a la tenperatura, por su patogenicidad en diferentes animales, por su especificidad hacia un determinado huésped, etc.. Cepas atenuadas aisladas en distintos sistemas virales han sido utilizadas como agentes vacunantes con resultados muy alentadores. Actualmente, se están empleando en Venezuela cepas de virus de la fiebre aftosa atenuadas para bovinos en la producción de vacunas. Estas cepas han sido utilizadas teniendo en cuenta su bajo grado de patogenicidad y alta calidad inmunogénica. Sin embargo, el criterio por el cual fueron seleccionadas es altamente empírico ya que se conoce muy poco acerca del mecanismo de atenuación. Principalmente, habian sido estudiadas en cuanto a sus caracteristicas biológicas de multiplicación respecto de las cepas virulentas. En base a lo previamente expuesto, se plantearon los siguientes objetivos para este tema de tesis: 1°) El estudio de los factores celulares y/o virales que influyen en el mecanismo de atenuación. 2°) Las caracteristicas inhibitorias de la multiplicación del virus salvaje por las cepas atenuadas. Estos objetivos fueron enfocados a traves de tres puntos principales: - Caracterización biológica del virus atenuado (infecciosidad, ciclo de multiplicación, etc.). - Caracterización de la interferencia por el virus atenuado en la multiplicación del virus salvaje. - Caracterización fisiquuímica del virus atenuado y de sus componentes (coeficiente de sedimentación, densidad, análisis de su genoma).Fil: Polacino, Patricia Susana. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina