African swine fever is a highly contagious viral disease of mandatory declaration to the World Organization for Animal Health (OIE). The lack of available vaccines makes its control difficult; thus, African swine fever virus (ASFV) represents a major threat to the swine industry. Inactivated vaccines do not confer solid protection against ASFV. Conversely, live attenuated viruses (LAV), either naturally isolated or obtained by genetic manipulation, have demonstrated reliable protection against homologous ASFV strains, although little or no protection has been demonstrated against heterologous viruses. Safety concerns are a major issue for the use of ASFV attenuated vaccine candidates and have hampered their implementation in the field so far. While trying to develop safer and efficient ASFV vaccines, we found that the deletion of the viral CD2v (EP402R) gene highly attenuated the virulent BA71 strain in vivo. Inoculation of pigs with the deletion mutant virus BA71ΔCD2 conferred protection not only against lethal challenge with the parental BA71 but also against the heterologous E75 (both genotype I strains). The protection induced was dose dependent, and the cross-protection observed in vivo correlated with the ability of BA71ΔCD2 to induce specific CD8+ T cells capable of recognizing both BA71 and E75 viruses in vitro. Interestingly, 100% of the pigs immunized with BA71ΔCD2 also survived lethal challenge with Georgia 2007/1, the genotype II strain of ASFV currently circulating in continental Europe. These results open new avenues to design ASFV cross-protective vaccines, essential to fight ASFV in areas where the virus is endemic and where multiple viruses are circulating.IMPORTANCE African swine fever virus (ASFV) remains enzootic in most countries of Sub-Saharan Africa, today representing a major threat for the development of their swine industry. The uncontrolled presence of ASFV has favored its periodic exportation to other countries, the last event being in Georgia in 2007. Since then, ASFV has spread toward neighboring countries, reaching the European Union's east border in 2014. The lack of available vaccines against ASFV makes its control difficult; so far, only live attenuated viruses have demonstrated solid protection against homologous experimental challenges, but they have failed at inducing solid cross-protective immunity against heterologous viruses. Here we describe a new LAV candidate with unique cross-protective abilities: BA71ΔCD2. Inoculation of BA71ΔCD2 protected pigs not only against experimental challenge with BA71, the virulent parental strain, but also against heterologous viruses, including Georgia 2007/1, the genotype II strain of ASFV currently circulating in Eastern Europe.

African swine fever is one of the most devastating pig diseases, against which there is no vaccine available. Recent work from our laboratory has demonstrated the protective potential of DNA vaccines encoding three African swine fever viral antigens (p54, p30, and the hemagglutinin extracellular domain) fused to ubiquitin. Partial protection was afforded in the absence of detectable antibodies prior to virus challenge, and survival correlated with the presence of a large number of hemagglutinin-specific CD8 IMPORTANCEAfrican swine fever is a highly contagious disease of domestic and wild pigs that is endemic in many sub-Saharan countries, where it causes important economic losses and is currently in continuous expansion across Europe. Unfortunately, there is no treatment nor an available vaccine. Early attempts using attenuated vaccines demonstrated their potential to protect pigs against experimental infection. However, their use in the field remains controversial due to safety issues. Although inactive and subunit vaccines did not confer solid protection against experimental ASFV infection, our DNA vaccination results have generated new expectations, confirming the key role of T-cell responses in protection and the existence of multiple ASFV antigens with protective potential, more of which are currently being identified. Thus, the future might bring complex and safe formulations containing more than a single viral determinant to obtain broadly protective vaccines. We believe that obtaining the optimal vaccine formulation it is just a matter of time, investment, and willingness.

African swine fever virus (ASFV) is the causal agent of African swine fever, a hemorrhagic and often lethal porcine disease causing enormous economical losses in affected countries. Endemic for decades in most of the sub-Saharan countries and Sardinia, the risk of ASFV-endemicity in Europe has increased since its last introduction into Europe in 2007. Live attenuated viruses have been demonstrated to induce very efficient protective immune responses, albeit most of the time protection was circumscribed to homologous ASFV challenges. However, their use in the field is still far from a reality, mainly due to safety concerns. In this study we compared the course of the in vivo infection caused by two homologous ASFV strains: the virulent E75 and the cell cultured adapted strain E75CV1, obtained from adapting E75 to grow in the CV1 cell-line. Interestingly, the kinetics of both viruses not only differed on the clinical signs that they caused and in the virus loads found, but also in the immunological pathways activated throughout the infections. Furthermore, E75CV1 confirmed its protective potential against the homologous E75 virus challenge and allowed the demonstration of poor cross-protection against BA71, thus defining it as heterologous. The in vitro specificity of the CD8+ T-cells present at the time of lethal challenge showed a clear activation against the homologous virus (E75) but not against BA71. These findings will be of utility for a better understanding of ASFV pathogenesis and for the rational designing of safe and efficient vaccines against this virus.

Early interactions of influenza A virus (IAV) with respiratory epithelium might determine the outcome of infection. The study of global cellular innate immune responses often masks multiple aspects of the mechanisms by which populations of cells work as organized and heterogeneous systems to defeat virus infection, and how the virus counteracts these systems. In this study, we experimentally dissected the dynamics of IAV and human epithelial respiratory cell interaction during early infection at the single-cell level. We found that the number of viruses infecting a cell (multiplicity of infection [MOI]) influences the magnitude of virus antagonism of the host innate antiviral response. Infections performed at high MOIs resulted in increased viral gene expression per cell and stronger antagonist effect than infections at low MOIs. In addition, single-cell patterns of expression of interferons (IFN) and IFN-stimulated genes (ISGs) provided important insights into the contributions of the infected and bystander cells to the innate immune responses during infection. Specifically, the expression of multiple ISGs was lower in infected than in bystander cells. In contrast with other IFNs, IFN lambda 1 (IFNL1) showed a widespread pattern of expression, suggesting a different cell-to-cell propagation mechanism more reliant on paracrine signaling. Finally, we measured the dynamics of the antiviral response in primary human epithelial cells, which highlighted the importance of early innate immune responses at inhibiting virus spread. IMPORTANCE Influenza A virus (IAV) is a respiratory pathogen of high importance to public health. Annual epidemics of seasonal IAV infections in humans are a significant public health and economic burden. IAV also causes sporadic pandemics, which can have devastating effects. The main target cells for IAV replication are epithelial cells in the respiratory epithelium. The cellular innate immune responses induced in these cells upon infection are critical for defense against the virus, and therefore, it is important to understand the complex interactions between the virus and the host cells. In this study, we investigated the innate immune response to IAV in the respiratory epithelium at the single-cell level, providing a better understanding on how a population of epithelial cells functions as a complex system to orchestrate the response to virus infection and how the virus counteracts this system.

African swine fever (ASF) is a pathology of pigs against which there is no treatment or vaccine. Understanding the equilibrium between innate and adaptive protective responses and immune pathology might contribute to the development of strategies against ASFV. Here we compare, using a proteomic approach, the course of the in vivo infection caused by two homologous strains: the virulent E75 and the attenuated E75CV1. Our results show a progressive loss of proteins by day 7 post-infection (pi) with E75, reflecting tissue destruction. Many signal pathways were affected by both infections but in different ways and extensions. Cytoskeletal remodelling and clathrin-endocytosis were affected by both isolates, while a greater number of proteins involved on inflammatory and immunological pathways were altered by E75CV1. 14-3-3 mediated signalling, related to immunity and apoptosis, was inhibited by both isolates. The implication of the Rho GTPases by E75CV1 throughout infection is also evident. Early events reflected the lack of E75 recognition by the immune system, an evasion strategy acquired by the virulent strains, and significant changes at 7 days post-infection (dpi), coinciding with the peak of infection and the time of death. The protein signature at day 31 pi with E75CV1 seems to reflect events observed at 1 dpi, including the upregulation of proteosomal subunits and molecules described as autoantigens (vimentin, HSPB1, enolase and lymphocyte cytosolic protein 1), which allow the speculation that auto-antibodies could contribute to chronic ASFV infections. Therefore, the use of proteomics could help understand ASFV pathogenesis and immune protection, opening new avenues for future research.Electronic supplementary materialThe online version of this article (10.1186/s13567-018-0585-z) contains supplementary material, which is available to authorized users.

Influenza A viruses (IAVs) cause seasonal epidemics which result in an important health and economic burden. Wild aquatic birds are the natural host of IAV. However, IAV can infect diverse hosts, including humans, domestic poultry, pigs, and others. IAVs circulating in animals occasionally cross the species barrier, infecting humans, which results in mild to very severe disease. In some cases, these viruses can acquire the ability to be transmitted among humans and initiate a pandemic. The nonstructural 1 (NS1) protein of IAV is an important antagonist of the innate immune response. In this study, using recombinant viruses and primary human cells, we show that NS1 proteins from human and avian hosts show intrinsic differences in the modulation of the innate immunity in human dendritic cells and epithelial cells, as well as different cellular localization dynamics in infected cells.

Background Leishmaniases are a group of neglected tropical parasitic diseases, mainly affecting vulnerable populations of countries with poor socioeconomic status. Development of efficient vaccines is a priority due to the increasing incidence of drug resistance and toxicity to current treatments. In the search for a safe and efficient protective vaccine for human and dog visceral leishmaniases, we analyzed the suitability of the immunomodulatory drug sirolimus (SIR) to boost a preventive DNA vaccine against leishmaniasis. SIR is an already marketed drug that has been described to boost immune protection against different disease models and has also emerged as a promising therapeutic drug against L. major. Methods Syrian hamsters were treated with SIR concomitantly with the administration of a DNA vaccine formulation consisting in four plasmids carrying the Leishmania genes LACK, TRYP, PAPLE22 and KMPII, respectively. Two weeks after the last vaccination, the animals were infected intraperitoneally with L. infantum parasites. Five weeks post-infection the parasite load was measured by real-time PCR in target tissues and immune response was evaluated by determining anti-Leishmania specific antibodies in combination with cytokine expression in the spleen. Results Our results show that the DNA vaccine itself efficiently reduced the burden of parasites in the skin (P = 0.0004) and lymph nodes (P = 0.0452). SIR administration also enhanced the protection by reducing the parasite load in the spleen (P = 0.0004). Vaccinated animals with or without SIR co-treatment showed lower IFN-γ expression levels than those found in the spleen of control animals. mRNA expression levels of NOS2 and IL-10 were found to be significantly higher in the vaccinated plus SIR treated group. Conclusions Co-administration of SIR enhances a DNA vaccination regimen against L. infantum, improving the reduction of parasite load in skin, lymph node and spleen. The analysis of immune markers in the spleen after challenge suggests that the trend to recover naïve levels of IFN-γ and IL-10, and the concurrent higher expression of NOS2, may be responsible for the protection induced by our vaccine co-administered with SIR.

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