Sofosbuvir shows a protective effect against vertical transmission of Zika virus and the associated congenital syndrome in rhesus monkeys.

The outbreaks of Zika virus (ZIKV) infection in Brazil, 2015-2016, were associated with severe congenital malformations. Our translational study aimed to test the efficacy of the antiviral agent sofosbuvir (SOF) against vertical transmission of ZIKV and the associated congenital syndrome (CZS), using a rhesus monkey model. Eight pregnant macaques were successfully infected during the organogenesis phase with a Brazilian ZIKV strain; five of them received SOF from two to fifteen days post-infection.

Purification of yellow fever virus produced in Vero cells for inactivated vaccine manufacture.

Yellow fever (YF) is a high-lethality viral disease, endemic in tropical regions of South America and Africa, with a population of over 900 million people under risk. A highly effective attenuated vaccine, produced in embryonated eggs, has been used for about 80 years. However, egg-based production limits manufacturing capacity, and vaccine shortage led to the emergency use of a fractional dose (1/5) by the WHO in an outbreak in Africa in 2016 and by Brazilian authorities during an outbreak in 2018.

Plant-Produced Subunit Vaccine Candidates against Yellow Fever Induce Virus Neutralizing Antibodies and Confer Protection against Viral Challenge in Animal Models.

Yellow fever (YF) is a viral disease transmitted by mosquitoes and endemic mostly in South America and Africa with 20-50% fatality. All current licensed YF vaccines, including YF-Vax (Sanofi-Pasteur, Lyon, France) and 17DD-YFV (Bio-Manguinhos, Rio de Janeiro, Brazil), are based on live attenuated virus produced in hens' eggs and have been widely used. The YF vaccines are considered safe and highly effective.

Heparin removal by ecteola-cellulose pre-treatment enables the use of plasma samples for accurate measurement of anti-Yellow fever virus neutralizing antibodies.

Technological innovations in vaccinology have recently contributed to bring about novel insights for the vaccine-induced immune response. While the current protocols that use peripheral blood samples may provide abundant data, a range of distinct components of whole blood samples are required and the different anticoagulant systems employed may impair some properties of the biological sample and interfere with functional assays. Although the interference of heparin in functional assays for viral neutralizing antibodies such as the functional plaque-reduction neutralization test (PRNT), considered the gold-standard method to assess and monitor the protective immunity induced by the Yellow fever virus (YFV) vaccine, has been well characterized, the development of pre-analytical treatments is still required for the establishment of optimized protocols.

CD4/CD8 Ratio and KT Ratio Predict Yellow Fever Vaccine Immunogenicity in HIV-Infected Patients.

Background: HIV-infected individuals have deficient responses to Yellow Fever vaccine (YFV) and may be at higher risk for adverse events (AE). Chronic immune activation-characterized by low CD4/CD8 ratio or high indoleamine 2,3-dioxygenase-1 (IDO) activity-may influence vaccine response in this population.

Methods: We prospectively assessed AE, viremia by the YFV virus and YF-specific neutralizing antibodies (NAb) in HIV-infected (CD4>350) and -uninfected adults through 1 year after vaccination.

CD4/CD8 Ratio Predicts Yellow Fever Vaccine-Induced Antibody Titers in Virologically Suppressed HIV-Infected Patients.

Background: Yellow fever vaccine (YFV) induces weaker immune responses in HIV-infected individuals. However, little is known about YFV responses among antiretroviral-treated patients and potential immunological predictors of YFV response in this population.

Methods: We enrolled 34 antiretroviral therapy (ART)-treated HIV-infected and 58 HIV-uninfected adults who received a single YFV dose to evaluate antibody levels and predictors of immunity, focusing on CD4(+) T-cell count, CD4(+)/CD8(+) ratio, and Human Pegivirus (GBV-C) viremia.

An inactivated yellow fever 17DD vaccine cultivated in Vero cell cultures.

Yellow fever is an acute infectious disease caused by prototype virus of the genus Flavivirus. It is endemic in Africa and South America where it represents a serious public health problem causing epidemics of hemorrhagic fever with mortality rates ranging from 20% to 50%. There is no available antiviral therapy and vaccination is the primary method of disease control.

Development of a membrane adsorber based capture step for the purification of yellow fever virus.

Yellow fever (YF) is an endemic disease in some tropical areas of South America and Africa that presents lethality rate between 20 and 50%. There is no specific treatment and to control this disease a highly effective live-attenuated egg based vaccine is widely used for travelers and residents of areas where YF is endemic. However, recent reports of rare, sometimes fatal, adverse events post-vaccination have raised concerns.

The synergistic effect of combined immunization with a DNA vaccine and chimeric yellow fever/dengue virus leads to strong protection against dengue.

The dengue envelope glycoprotein (E) is the major component of virion surface and its ectodomain is composed of domains I, II and III. This protein is the main target for the development of a dengue vaccine with induction of neutralizing antibodies. In the present work, we tested two different vaccination strategies, with combined immunizations in a prime/booster regimen or simultaneous inoculation with a DNA vaccine (pE1D2) and a chimeric yellow fever/dengue 2 virus (YF17D-D2).

17DD yellow fever vaccine: a double blind, randomized clinical trial of immunogenicity and safety on a dose-response study.

Objective: To verify if the Bio-Manguinhos 17DD yellow fever vaccine (17DD-YFV) used in lower doses is as immunogenic and safe as the current formulation.

Results: Doses from 27,476 IU to 587 IU induced similar seroconversion rates and neutralizing antibodies geometric mean titers (GMTs). Immunity of those who seroconverted to YF was maintained for 10 mo.

Induction of a protective response in mice by the dengue virus NS3 protein using DNA vaccines.

The dengue non-structural 3 (NS3) is a multifunctional protein, containing a serino-protease domain, located at the N-terminal portion, and helicase, NTPase and RTPase domains present in the C-terminal region. This protein is considered the main target for CD4+ and CD8+ T cell responses during dengue infection, which may be involved in protection. However, few studies have been undertaken evaluating the use of this protein as a protective antigen against dengue, as well as other flavivirus.

DNA vaccines against dengue virus type 2 based on truncate envelope protein or its domain III.

Two DNA vaccines were constructed encoding the ectodomain (domains I, II and III) of the DENV2 envelope protein (pE1D2) or only its domain III (pE2D2), fused to the human tissue plasminogen activator signal peptide (t-PA). The expression and secretion of recombinant proteins was confirmed in vitro in BHK cells transfected with the two plasmids, detected by immunofluorescence or immunoprecipitation of metabolically labeled gene products, using polyclonal and monoclonal antibodies against DENV2. Besides, results reveal that the ectodomain of the E protein can be efficiently expressed in vivo, in a mammalian system, without the prM protein that is hypothesized to act as a chaperonin during dengue infection.

Effects of hydrostatic pressure on the stability and thermostability of poliovirus: a new method for vaccine preservation.

Viruses are a structurally diverse group of infectious agents that differ widely in their sensitivities to high hydrostatic pressure (HHP). Studies on picornaviruses have demonstrated that these viruses are extremely resistant to HHP treatments, with poliovirus appearing to be the most resistant. Here, the three attenuated poliovirus serotypes were compared with regard to pressure and thermal resistance.

Production of yellow fever virus in microcarrier-based Vero cell cultures.

In this work, the propagation of the 17DD yellow fever virus in Vero cells grown on Cytodex-1 microcarriers was evaluated. After verifying that upon infection the virus adsorption step could be performed under continuous agitation, experiments were carried out in spinners and sparged lab-scale stirred-tank bioreactor to evaluate the use of a commercial serum-free medium (VP-SFM) and to investigate the effects of multiplicity of infection (MOI) and time of infection (TOI) on virus production. Virus titers as high as 8.

Risk factors for dengue virus infection in rural Amazonia: population-based cross-sectional surveys.

A comparison of dengue virus (DENV) antibody levels in paired serum samples collected from predominantly DENV-naive residents in an agricultural settlement in Brazilian Amazonia (baseline seroprevalence, 18.3%) showed a seroconversion rate of 3.67 episodes/100 person-years at risk during 12 months of follow-up.

Limited replication of yellow fever 17DD and 17D-Dengue recombinant viruses in rhesus monkeys.

For the development of safe live attenuated flavivirus vaccines one of the main properties to be established is viral replication. We have used real-time reverse transcriptase-polymerase chain reaction and virus titration by plaque assay to determine the replication of yellow fever 17DD virus (YFV 17DD) and recombinant yellow fever 17D viruses expressing envelope proteins of dengue virus serotypes 2 and 4 (17D-DENV-2 and 17D-DENV-4). Serum samples from rhesus monkeys inoculated with YFV 17DD and 17D-DENV chimeras by intracerebral or subcutaneous route were used to determine and compare the viremia induced by these viruses.

Pressure-inactivated yellow fever 17DD virus: implications for vaccine development.

The successful Yellow Fever (YF) vaccine consists of the live attenuated 17D-204 or 17DD viruses. Despite its excellent record of efficacy and safety, serious adverse events have been recorded and influenced extensive vaccination in endemic areas. Therefore, alternative strategies should be considered, which may include inactivated whole virus.

Study of the genetic stability of measles virus CAM-70 vaccine strain after serial passages in chicken embryo fibroblasts primary cultures.

To evaluate the genetic stability of the CAM-70 measles vaccine strain we have performed 10 serial passages of the seed lot virus FMS-7 in chicken embryo fibroblasts primary cultures (CEF) under production conditions. The nucleotide sequences of the seed lot virus, the virus from a vaccine vial (third passage) and from the 10th passage were determined and compared with each other and with sequences from other sources. The full genome analysis of the CAM-70 vaccine still considers it as the most divergent among all vaccine strains.

Construction and characterization of recombinant flaviviruses bearing insertions between E and NS1 genes.

Background: The yellow fever virus, a member of the genus Flavivirus, is an arthropod-borne pathogen causing severe disease in humans. The attenuated yellow fever 17D virus strain has been used for human vaccination for 70 years and has several characteristics that are desirable for the development of new, live attenuated vaccines. We described here a methodology to construct a viable, and immunogenic recombinant yellow fever 17D virus expressing a green fluorescent protein variant (EGFP).

Expression of foreign protein epitopes at the surface of recombinant yellow fever 17D viruses based on three-dimensional modeling of its envelope protein.

The yellow fever (YF) 17D vaccine is a live attenuated virus, and its genetic manipulation constitutes a new platform for vaccine development. In this article, we review one of the possible approaches to enable this development, which is the insertion of foreign protein epitopes into different locations of the genome. We describe the three-dimensional (3D) modeling of the YF 17D virus E protein structure based on tick-borne encephalitis (TBE) and the identification of a potential insertion site located at the YF 17D fg loop.

Yellow fever vaccination: some thoughts on how much is enough [Vaccine 23 (2005) 3908-3914].

In a recently published article in this journal, Massad et al. contraindicates yellow fever vaccination to persons 60 years or older, considering that the risk of serious adverse events is higher for this age class. The conclusion was based on the input of available data on age-related probabilities of developing serious adverse events in the United States, as well on other data not firmly established.

Molecular and phenotypic analysis of a working seed lot of yellow fever virus 17DD vaccine strain produced from the secondary seed lot 102/84 with an additional passage in chicken embryos.

Over the last 17 years, the yellow fever (YF) 17DD vaccine secondary seed lot 102/84 was used to produce many million doses of vaccine but it was recently used up. In the absence of other lots at the same passage level a large vaccine batch produced from 102/84 was turned into a new working seed. This new seed was characterized with regard to attenuation in the recommended internationally accepted monkey neurovirulence test (MNVT) using the 102/84 virus as reference.

Protection against dengue type 2 virus induced in mice immunized with a DNA plasmid encoding the non-structural 1 (NS1) gene fused to the tissue plasminogen activator signal sequence.

Dengue is one of the most important arboviral diseases in humans, and although efforts over the last decades have dealt with the development of a vaccine, this vaccine is not available yet. In order to evaluate the potential of a DNA vaccine based on the non-structural 1 (NS1) protein against dengue virus (DENV), we constructed the pcTPANS1 plasmid which contains the secretory signal sequence derived from human tissue plasminogen activator (t-PA) fused to the full length of the DENV-2 NS1 gene. Results indicate that pcTPANS1 promotes correct expression of NS1 in eukaryotic cells and drives secretion of the recombinant protein to the surrounding medium in a dimeric form.

Attenuation of recombinant yellow fever 17D viruses expressing foreign protein epitopes at the surface.

The yellow fever (YF) 17D vaccine is a live attenuated virus. Three-dimensional (3D) homology modeling of the E protein structure from YF 17D virus and its comparison with that from tick-borne encephalitis virus revealed that it is possible to accommodate inserts of different sizes and amino acid compositions in the flavivirus E protein fg loop. This is consistent with the 3D structures of both the dimeric and trimeric forms in which the fg loop lies exposed to solvents.

Production of yellow fever 17DD vaccine virus in primary culture of chicken embryo fibroblasts: yields, thermo and genetic stability, attenuation and immunogenicity.

While a good vaccine against yellow fever (YF) virus has been available for decades, the basic technology for the production of YF vaccine in chicken embryos has remained substantially unchanged since the 1940s. Here we describe the highly efficient and economic production of the 17DD strain of YF virus in chicken embryo fibroblast (CEF) cell cultures with viral titers ranging from 6.3 to 6.