The role of matrix protein 2 ectodomain in the development of universal influenza vaccines

The influenza A virus matrix protein 2 ectodomain (M2e) is a universal influenza A vaccine candidate. Numerous studies in laboratory mice, but very few in natural influenza A virus hosts, have demonstrated that M2e-based vaccines can provide protection against any influenza A virus challenge. M2e-based immunity is largely accomplished by IgG and early stage clinical studies have demonstrated that the vaccine is safe. Yet M2e is considered a difficult target to develop as a vaccine: it does not offer sterilizing immunity and its mode of action relies on Fc. receptor-mediated effector mechanisms, most likely in concert with alveolar macrophages. In a human challenge study with an H3N2 virus, treatment with a monoclonal M2e-specific human IgG was associated with a faster recovery compared to placebo treatment. If the universal influenza vaccine field incorporates this antigen into next generation vaccines, M2e could prove its merit when the next influenza pandemic strikes

Cell-mediated responses to human metapneumovirus infection

Viruses are the most common cause of acute respiratory tract infections (ARTI). Human metapneumovirus (hMPV) frequently causes viral pneumonia which can become life-threatening if the virus spreads to the lungs. Even though hMPV was only isolated in 2001, this negative-stranded RNA virus has probably been circulating in the human population for many decades. Interestingly, almost all adults have serologic evidence of hMPV infection. A well-established host immune response is evoked when hMPV infection occurs. However, the virus has evolved to circumvent and even exploit the host immune response. Further, infection with hMPV induces a weak memory response, and re-infections during life are common. In this review, we provide a comprehensive overview of the different cell types involved in the immune response in order to better understand the immunopathology induced by hMPV. Such knowledge may contribute to the development of vaccines and therapeutics directed against hMPV

Single-domain antibodies and their formatting to combat viral infections

Since their discovery in the 1990s, single-domain antibodies (VHHs), also known as NanobodiesA (R), have changed the landscape of affinity reagents. The outstanding solubility, stability, and specificity of VHHs, as well as their small size, ease of production and formatting flexibility favor VHHs over conventional antibody formats for many applications. The exceptional ease by which it is possible to fuse VHHs with different molecular modules has been particularly explored in the context of viral infections. In this review, we focus on VHH formats that have been developed to combat viruses including influenza viruses, human immunodeficiency virus-1 (HIV-1), and human respiratory syncytial virus (RSV). Such formats may significantly increase the affinity, half-life, breadth of protection of an antiviral VHH and reduce the risk of viral escape. In addition, VHHs can be equipped with effector functions, for example to guide components of the immune system with high precision to sites of viral infection

TIV vaccination modulates host responses to influenza virus infection that correlate with protection against bacterial superinfection

Background: Influenza virus infection predisposes to secondary bacterial pneumonia. Currently licensed influenza vaccines aim at the induction of neutralizing antibodies and are less effective if the induction of neutralizing antibodies is low and/or the influenza virus changes its antigenic surface. We investigated the effect of suboptimal vaccination on the outcome of post-influenza bacterial superinfection. Methods: We established a mouse vaccination model that allows control of disease severity after influenza virus infection despite inefficient induction of virus-neutralizing antibody titers by vaccination. We investigated the effect of vaccination on virus-induced host immune responses and on the outcome of superinfection with Staphylococcus aureus. Results: Vaccination with trivalent inactivated virus vaccine (TIV) reduced morbidity after influenza A virus infection but did not prevent virus replication completely. Despite the poor induction of influenza-specific antibodies, TIV protected from mortality after bacterial superinfection. Vaccination limited loss of alveolar macrophages and reduced levels of infiltrating pulmonary monocytes after influenza virus infection. Interestingly, TIV vaccination resulted in enhanced levels of eosinophils after influenza virus infection and recruitment of neutrophils in both lungs and mediastinal lymph nodes after bacterial superinfection. Conclusion: These observations highlight the importance of disease modulation by influenza vaccination, even when suboptimal, and suggest that influenza vaccination is still beneficial to protect during bacterial superinfection in the absence of complete virus neutralization

An inside job : applications of intracellular single domain antibodies

Sera of camelid species contain a special kind of antibody that consists only of heavy chains. The variable antigen binding domain of these heavy chain antibodies can be expressed as a separate entity, called a single domain antibody that is characterized by its small size, high solubility and oftentimes exceptional stability. Because of this, most single domain antibodies fold correctly when expressed in the reducing environment of the cytoplasm, and thereby retain their antigen binding specificity. Single domain antibodies can thus be used to target a broad range of intracellular proteins. Such intracellular single domain antibodies are also known as intrabodies, and have proven to be highly useful tools for basic research by allowing visualization, disruption and even targeted degradation of intracellular proteins. Furthermore, intrabodies can be used to uncover prospective new therapeutic targets and have the potential to be applied in therapeutic settings in the future. In this review we provide a brief overview of recent advances in the field of intracellular single domain antibodies, focusing on their use as research tools and potential therapeutic applications. Special attention is given to the available methods that allow delivery of single domain antibodies into cells

Respiratory syncytial virus nonstructural proteins 1 and 2 : exceptional disrupters of innate immune responses

Human respiratory syncytial virus (RSV) is the most important cause of acute lower respiratory tract disease in infants worldwide. As a first line of defense against respiratory infections, innate immune responses, including the production of type I and III interferons (IFNs), play an important role. Upon infection with RSV, multiple pattern recognition receptors (PRRs) can recognize RSV-derived pathogen-associated molecular patterns (PAMPs) and mount innate immune responses. Retinoic-acid-inducible gene-I (RIG-I) and nucleotide-binding oligomerization domain-containing protein 2 (NOD2) have been identified as important innate receptors to mount type I IFNs during RSV infection. However, type I IFN levels remain surprisingly low during RSV infection despite strong viral replication. The poor induction of type I IFNs can be attributed to the cooperative activity of 2 unique, nonstructural (NS) proteins of RSV, i.e., NS1 and NS2. These viral proteins have been shown to suppress both the production and signaling of type I and III IFNs by counteracting a plethora of key host innate signaling proteins. Moreover, increasing numbers of IFN-stimulated genes (ISGs) are being identified as targets of the NS proteins in recent years, highlighting an underexplored protein family in the identification of NS target proteins. To understand the diverse effector functions of NS1 and NS2, Goswami and colleagues proposed the hypothesis of the NS degradasome (NSD) complex, a multiprotein complex made up of, at least, NS1 and NS2. Furthermore, the crystal structure of NS1 was resolved recently and, remarkably, identified NS1 as a structural paralogue of the RSV matrix protein. Unfortunately, no structural data on NS2 have been published so far. In this review, we briefly describe the PRRs that mount innate immune responses upon RSV infection and provide an overview of the various effector functions of NS1 and NS2. Furthermore, we discuss the ubiquitination effector functions of NS1 and NS2, which are in line with the hypothesis that the NSD shares features with the canonical 26S proteasome

Illumina MiSeq sequencing disfavours a sequence motif in the GFP reporter gene

Green fluorescent protein (GFP) is one of the most used reporter genes. We have used next-generation sequencing (NGS) to analyse the genetic diversity of a recombinant influenza A virus that expresses GFP and found a remarkable coverage dip in the GFP coding sequence. This coverage dip was present when virus-derived RT-PCR product or the parental plasmid DNA was used as starting material for NGS and regardless of whether Nextera XT transposase or Covaris shearing was used for DNA fragmentation. Therefore, the sequence coverage dip in the GFP coding sequence was not the result of emerging GFP mutant viruses or a bias introduced by Nextera XT fragmentation. Instead, we found that the Illumina MiSeq sequencing method disfavours the 'CCCGCC' motif in the GFP coding sequence

Controlling Influenza by Cytotoxic T-Cells: Calling for Help from Destroyers

Influenza is a vaccine preventable disease that causes severe illness and excess mortality in humans. Licensed influenza vaccines induce humoral immunity and protect against strains that antigenically match the major antigenic components of the vaccine, but much less against antigenically diverse influenza strains. A vaccine that protects against different influenza viruses belonging to the same subtype or even against viruses belonging to more than one subtype would be a major advance in our battle against influenza. Heterosubtypic immunity could be obtained by cytotoxic T-cell (CTL) responses against conserved influenza virus epitopes. The molecular mechanisms involved in inducing protective CTL responses are discussed here. We also focus on CTL vaccine design and point to the importance of immune-related databases and immunoinformatics tools in the quest for new vaccine candidates. Some techniques for analysis of T-cell responses are also highlighted, as they allow estimation of cellular immune responses induced by vaccine preparations and can provide correlates of protection