ABSTRACT
Here we report on new subunit vaccines based on recombinant yeast of the type Kluyveromyces lactis (K. lactis), which protect mice from a lethal influenza A virus infection. Applying a genetic system that enables the rapid generation of transgenic yeast, we have developed K. lactis strains that express the influenza A virus hemagglutinin, HA, either individually or in combination with the viral M1 matrix protein. Subcutaneous application of the inactivated, but otherwise non-processed yeast material shows a complete protection of BALB/c mice in prime/boost and even one-shot/single dose vaccination schemes against a subsequent, lethal challenge with the cognate influenza virus. The yeast vaccines induce titers of neutralizing antibodies that are readily comparable to those induced by an inactivated virus vaccine. These data suggest that HA and M1 are produced with a high antigenicity in the yeast cells. Based on these findings, multivalent, DIVA-capable, yeast-based subunit vaccines may be developed as promising alternatives to conventional virus-based anti-flu vaccines for veterinary applications.
Subject(s)
Influenza A virus/immunology , Influenza Vaccines/immunology , Influenza, Human/prevention & control , Vaccination , Vaccines, DNA/immunology , Vaccines, Subunit/immunology , Animals , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Disease Models, Animal , Gene Expression , Hemagglutinin Glycoproteins, Influenza Virus/genetics , Hemagglutinin Glycoproteins, Influenza Virus/immunology , Humans , Immunization Schedule , Influenza Vaccines/administration & dosage , Influenza Vaccines/genetics , Mice , Vaccination/methods , Vaccines, DNA/administration & dosage , Vaccines, DNA/genetics , Vaccines, Subunit/administration & dosage , Vaccines, Subunit/genetics , Yeasts/genetics , Yeasts/immunologyABSTRACT
Potent adjuvant systems are required for subunit and single antigen based vaccines to provide sufficient immunogenicity. Furthermore, adjuvants can reduce the required number of immunisations and the antigen amount. Squalene nanoemulsions, like MF59® and AddaVax™, are potent, safe and well characterised adjuvant systems and approved for use in humans. Here, we developed squalene containing solid lipid nanoparticles, which can be sterilised by steam sterilisation and stored as freeze-dried power together with a yeast-based vaccine. Detailed size measurements using dynamic and static light scattering were applied, as the immune stimulating effect of squalene emulsions is mainly dependent on the particle size. The size range of AddaVax™ (120-170â¯nm) was favoured for the developed squalene containing solid lipid nanoparticles. Differential scanning calorimetry (DSC) and H NMR studies were performed to characterise the interactions of the incorporated liquid squalene with the solid hard fat matrix. A homogeneous distribution as liquid domains in the solid glyceride structure was suggested for the liquid squalene. The developed adjuvant was compared with Freund's adjuvant and a commercially available squalene nanoemulsion in a vaccine trial in the mouse model with a yeast-based vaccine directed against the infectious bursal disease virus. All squalene-based adjuvants showed excellent biocompatibility and provided immune stimulating properties comparable to Freund's adjuvant.
Subject(s)
Adjuvants, Immunologic/chemistry , Freund's Adjuvant/chemistry , Lipids/chemistry , Nanoparticles/chemistry , Squalene/chemistry , Vaccines/chemistry , Yeast, Dried/chemistry , Animals , Birnaviridae Infections/immunology , Emulsions/chemistry , Female , Freund's Adjuvant/immunology , Immunization/methods , Infectious bursal disease virus/immunology , Lipids/immunology , Mice , Mice, Inbred BALB C , Particle Size , Squalene/immunology , Vaccines/immunology , Yeast, Dried/immunologyABSTRACT
The major obstacle to cure infections with human immunodeficiency virus (HIV-1) is integrated proviral genomes, which are not eliminated by antiretroviral therapies (ART). Treatment approaches with latency-reversing agents (LRAs) aim at inducing provirus expression to tag latently-infected cells for clearance through viral cytopathic effects or cytotoxic T cell (CTL) responses. However, the currently tested LRAs reveal evident drawbacks as gene expression is globally induced and viral outgrowth is insecure. Here, we present transcription activator-like effector (TALE) proteins as potent tools to activate HIV-1 specifically. The large variety of circulating HIV-1 strains and, accordingly, integrated proviruses was addressed by the programmable DNA-specificity of TALEs. Using customized engineered TALEs, a substantial transcription activation and viral outgrowth was achieved with cells obtained from different HIV-1 patients. Our data suggest that TALEs may be useful tools in future strategies aimed at removing HIV-1 reservoirs.
Subject(s)
HIV Infections/metabolism , HIV-1/physiology , Transcription Factors/metabolism , Virus Activation , Gene Expression Regulation, Viral , HIV Infections/genetics , HIV Infections/virology , HIV-1/genetics , Humans , Multigene Family , Species Specificity , Transcription Factors/genetics , Viral Proteins/genetics , Viral Proteins/metabolism , Virus LatencyABSTRACT
Infections with bovine viral diarrhea virus (BVDV) have a huge economic impact on cattle production and reproduction worldwide. A key factor for BVDV surveillance and eventual eradication is to efficiently detect infections and to monitor herd immunity. In this study, we generated a stable, bi-cistronic BVDV that encoded EGFP in addition to the viral proteins. Applying this recombinant virus, a new flow-cytometry-based virus neutralization test was established that enabled accurate and reliable detection of field-virus-infected and vaccinated animals. The test, which is simple and fast, is expected to support novel, effective screening procedures in eradication and vaccination programmes.