Pam3CSK4 adjuvant given intranasally boosts anti-Leishmania immunogenicity but not protective immune responses conferred by LaAg vaccine against visceral leishmaniasis

The use of adjuvants in vaccine formulations is a well-established practice to improve immunogenicity and protective immunity against diseases. Previously, we have demonstrated the feasibility of intranasal vaccination with the antigen of killed Leishmania amazonensis promastigotes (LaAg) against experimental leishmaniasis. In this work, we sought to optimize the immunogenic effect and protective immunity against murine visceral leishmaniasis conferred by intranasal delivery of LaAg in combination with a synthetic TLR1/TLR2 agonist (Pam3CSK4). Intranasal vaccination with LaAg/PAM did not show toxicity or adverse effects, induced the increase of delayed-type hypersensitivity response and the production of inflammatory cytokines after parasite antigen recall. However, mice vaccinated with LaAg/PAM and challenged with Leishmania infantum presented significant reduction of parasite burden in both liver and spleen, similar to those vaccinated with LaAg. Although LaAg/PAM intranasal vaccination had induced higher frequencies of specific CD4⁺ and CD8⁺ T cells and increased levels of IgG2a antibody isotype in serum, both LaAg and LaAg/PAM groups presented similar levels of IL-4 and IFN-y and decreased production of IL-10 when compared to controls. Our results provide the first evidence of the feasibility of intranasal immunization with antigens of killed Leishmania in association with a TLR agonist, which may be explored for developing an effective and alternative strategy for vaccination against visceral leishmaniasis.     

Maternal Immunization Confers Protection to the Offspring against an Attaching and Effacing Pathogen through Delivery of IgG in Breast Milk

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Biotechnology of virus eradication and plant vaccination in phytobiome context

A plant’s associated biota plays an integral role in its metabolism, nutrient uptake, stress tolerance, pathogen resistance and other physiological processes. Although a virome is an integral part of the phytobiome, a major contradiction exists between the holobiont approach and the practical need to eradicate pathogens from agricultural crops. In this review, we discuss grapevine virus control, but the issue is also relevant for numerous other crops, including potato, cassava, citrus, cacao and other species. Grapevine diseases, especially viral infections, cause main crop losses. Methods have been developed to eliminate viruses and other microorganisms from plant material, but elimination of viruses from plant material does not guarantee protection from future reinfection. Elimination of viral particles in plant material could create genetic drift, leading in turn to an increase in the occurrence of pathogenic strains of viruses. A possible solution may be a combination of virus elimination and plant propagation in tissue culture with in vitro vaccination. In this context, possible strategies to control viral infections include application of plant resistance inducers, cross protection and vaccination using siRNA, dsRNA and viral replicons during plant ‘cleaning’ and in vitro propagation. The experience and knowledge accumulated in human immunization can help plant scientists to develop and employ new methods of protection, leading to more sustainable and healthier crop production.     

Virus-related Knowledge in Covid-19 Times - Results from two Cross-sectional Studies in Austria and Implications for School

Viruses have become a prominent issue in public health, politics and economics due to the Covid-19 pandemic. Yet they have still met little attention in educational research, although misconceptions concerning viruses may contribute to antibiotics misuse, disbelief in existence of viruses and distrust in vaccination. We investigated knowledge and attitudes in Austria concerning Covid-19, viruses in general and vaccination. We conducted two cross-sectional online surveys. Study A was performed Austrian-wide (N = 1027), study B specifically targeted Austrian students from middle and high schools (N = 1728). Several participants did not believe in the existence of SARS-CoV-2. General vaccination damage was highly overrated. Many defined viruses as unicellular organisms or bacteria, and 6-10 % believed that they can be killed by antibiotics. Very many participants were unable to identify, whether a specific disease was caused by a virus or another pathogen. Knowledge was significantly correlated with level of education/grade and interest in virology. Additionally, willingness to become vaccinated was significantly correlated with knowledge. Many participants felt insufficiently informed about viruses at school. We strongly recommend that virus-related school education must highly improve to enable the population to correctly assess health-related information, counter fake news and come to scientifically informed decisions.     

Strategies for antigen choice and priming of dendritic cells influence the polarization and efficacy of antitumor T-cell responses in dendritic cell–based cancer vaccination

Dendritic cells (DCs) primed with tumor antigens (Ags) can stimulate tumor rejection. This study was aimed at evaluating the polarization of T-cell responses using various DC Ag-priming strategies for vaccination purposes. DCs cocultured with irradiated apoptotic tumor cells, DC-tumor fusions, and DCs pulsed with freeze-thaw tumor lysate Ags served as Ag-primed DCs, with EG7 tumor cells (class II negative) expressing OVA as the model Ag. DCs loaded with class I– and class II–restricted OVA synthetic peptides served as controls. Primed DCs were assessed by the in vitro activation of B3Z OVA-specific CD8 T cells and the proliferation of OVA-specific CD8 and CD4 T cells from OT-I and OT-II TCR transgenic mice, respectively. In vivo responses were measured by tumor regression following treatment with Ag-primed DCs and by CTL assays. Quantification of IL-2, IL-4, IL-5, IFN-, and TNF- by cytometric bead array (CBA) assay determined the polarization of TH1/TH2 responses, whereas H-2 K^b /SIINFEKL tetramers monitored the expansion of OVA-specific T cells. DC-EG7 hybrids stimulated both efficient class I and class II OVA responses, showing that DC-tumor hybrids are also capable of class II cross-presentation. The hybrids also induced the most potent CTLs, offered the highest protection against established EG7 tumors and also induced the highest stimulation of IFN- and TNF- production. DCs cocultured with irradiated EG7 were also effective at inducing OVA-specific responses, however with slightly reduced potency to those evoked by the hybrids. DCs loaded with lysates Ags were much less efficient at stimulating any of the OVA-specific T-cell responses, showed very little antitumor protection, and stimulated a weak TH1 response, overbalanced by an IL-5 TH2 response. The strategy of Ag-loading clearly influences the ability of DCs to polarize T cells for a TH1/TH2 response and thus determines the outcome of the elicited immune response, during various vaccination protocols.Abbreviations DC Dendritic cell - FSC Forward scatter - SSC Side scatter - TC Tumor cells This work was supported by Grant 9853 from the Leukaemia Research Fund, UK; a JRC studentship from GKT; and the Lewis Family Research Trust     

Immunization with a plant-produced colorectal cancer antigen

Cancer vaccination has become an important focus of oncology in recent years. Active immunization with tumor-associated antigens such as colorectal cancer antigen GA733-2 is thought to potentially overcome the reoccurrence of metastasis. As recombinant protein production in bioreactors is costly and subject to growing safety concerns, we tested plants as an alternative for the expression of a potential colorectal cancer vaccine. Comparing colorectal cancer antigen GA733-2 produced in tobacco plants with the same antigen produced in insect cell culture, we found a similar humoral immune response to injection of either of the two antigen preparations into mice. Some minor differences were observed in the cellular response that might be due to impurities. Our studies compare for the first time, immunization with the same antigen expressed in either plants or insect cell culture. This will provide important data for use of plants as production systems of therapeutics.     

Anti-idiotype responses abrogate anti-CD4–induced tolerance to a tumor-specific antigen and promote systemic tumor immunity

Purpose: Immunologic-based cancer treatment modalities represent an active area of investigation. Included in these strategies are passive administration of monoclonal antibodies which recognize tumor-associated antigens and active vaccination with identified tumor antigens. However, several problems associated with these types of treatment strategies have been identified. Methods: In this report, we address certain issues by employing a murine model for experimental pulmonary metastasis and a tumor antigen vaccination strategy that induces complete tumor immunity in this system. Utilizing this model, we attempt to address issues related to unresponsiveness to tumor antigen immunization induced by passive administration of a rat monoclonal anti-CD4 and the induction of anti-idiotype responses to a passively administered monoclonal antibody and the effects on the induction of tumor immunity. Results: The results presented indicate that passive administration of rat monoclonal anti-CD4 exhibits immunosuppressive effects that inhibit the production of antibodies to the tumor antigen immunization and abolishes tumor immunity. Repeated administration of the rat monoclonal anti-CD4 results in an anti-idiotype response that can abrogate unresponsiveness to tumor antigen immunization and promote systemic tumor immunity. Conclusions: The data examine a number of potential problems associated with immunologic-based treatments for cancer. These problems include the potential for tolerance to the tumor antigen and establishing an immunocompromised state where immunization with a tumor antigen failed to generate tumor immunity. Approaches to eliminate tolerant T cells by targeting anti-CD4 via anti-idiotype responses that could be generated in vivo without CD4⁺ T cells allowed for recovery of nontolerant T cells, and an antibody response to the tumor antigen that results in tumor immunity.Abbreviations CTL Cytotoxic T lymphocyte - FITC Fluorescein isothiocyanate - OD Optical density - PBS Phosphate-buffered saline - SV40 Simian virus 40     

Bacteriophage-mediated nucleic acid immunisation

Whole bacteriophage lambda particles, containing reporter genes under the control of the cytomegalovirus promoter (P(CMV)), have been used as delivery vehicles for nucleic acid immunisation. Following intramuscular injection of mice with lambda-gt11 containing the gene for hepatitis B surface antigen (HBsAg), anti-HBsAg responses in excess of 150 mIU ml(-1) were detected. When isolated peritoneal macrophages were incubated with whole lambda particles containing the gene for green fluorescent protein (GFP) under the control of P(CMV), GFP antigen was detected on the macrophage surface 8 h later. Results suggested that direct targeting of antigen-presenting cells by bacteriophage 'vaccines' may occur, leading to enhanced immune responses compared to naked DNA delivery. Bacteriophage DNA vaccines offer several advantages: they do not contain antibiotic resistance genes, they offer a large cloning capacity (approximately 15 kb), the DNA is protected from environmental degradation, they offer the potential for oral delivery, and large-scale production is cheap, easy and extremely rapid.