The SARS-CoV-2 spike (S) glycoprotein is synthesized as a large precursor protein and must be activated by proteolytic cleavage into S1 and S2. A recombinant modified vaccinia virus Ankara (MVA) expressing native, full-length S protein (MVA-SARS-2-S) is currently under investigation as a candidate vaccine in phase I clinical studies. Initial results from immunogenicity monitoring revealed induction of S-specific antibodies binding to S2, but low-level antibody responses to the S1 domain. Follow-up investigations of native S antigen synthesis in MVA-SARS-2-S–infected cells revealed limited levels of S1 protein on the cell surface. In contrast, we found superior S1 cell surface presentation upon infection with a recombinant MVA expressing a stabilized version of SARS-CoV-2 S protein with an inactivated S1/S2 cleavage site and K986P and V987P mutations (MVA-SARS-2-ST). When comparing immunogenicity of MVA vector vaccines, mice vaccinated with MVA-SARS-2-ST mounted substantial levels of broadly reactive anti-S antibodies that effectively neutralized different SARS-CoV-2 variants. Importantly, intramuscular MVA-SARS-2-ST immunization of hamsters and mice resulted in potent immune responses upon challenge infection and protected from disease and severe lung pathology. Our results suggest that MVA-SARS-2-ST represents an improved clinical candidate vaccine and that the presence of plasma membrane–bound S1 is highly beneficial to induce protective antibody levels.

The rapid development and deployment of injectable coronavirus disease 2019 (COVID-19) vaccines - in combination with non-pharmaceutical interventions and development of treatment options - significantly contributed to a decrease in both infection and mortality rates during the pandemic and saved millions of lives. However, injectable vaccines do not robustly and consistently induce a mucosal immune response, which is considered a key factor to prevent infection with and transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Hence, a tremendous effort is being made globally to develop next generation COVID-19 vaccines, which are capable of inducing a robust mucosal immune response in addition to a strong systemic cellular and humoral immune response. Mucosal COVID-19 vaccines have been evaluated successfully in preclinical and clinical trials, in which protection against SARS-CoV-2 infection has been demonstrated. This protective efficacy was associated with the upregulation of secretory IgA antibodies and the maturation of tissue-resident memory cells in the respiratory tract, which, together with an induced systemic immune response, significantly reduced viral replication and transmission in animal models. However, only five active mucosal vaccines (plus one 'passive' vaccine) have received approval for human use and robust data on their efficacy in inducing mucosal immune responses in humans and in blocking infection and transmission are missing. This highlights the importance of expanded research in this field. In this review, we aim to summarize what is known about these currently licensed vaccines, with an emphasis on the key findings obtained in both preclinical and clinical studies.

Influenza, a respiratory disease mainly caused by influenza A and B, viruses of the Orthomyxoviridae, is still a burden on our society’s health and economic system. Influenza A viruses (IAV) circulate in mammalian and avian populations, causing seasonal outbreaks with high numbers of cases. Due to the high variability in seasonal IAV triggered by antigenic drift, annual vaccination is necessary, highlighting the need for a more broadly protective vaccine against IAV. The safety tested Modified Vaccinia virus Ankara (MVA) is licensed as a third-generation vaccine against smallpox and serves as a potent vector system for the development of new candidate vaccines against different pathogens. Here, we generated and characterized recombinant MVA candidate vaccines that deliver the highly conserved internal nucleoprotein (NP) of IAV under the transcriptional control of five newly designed chimeric poxviral promoters to further increase the immunogenic properties of the recombinant viruses (MVA-NP). Infections of avian cell cultures with the recombinant MVA-NPs demonstrated efficient synthesis of the IAV-NP which was expressed under the control of the five new promoters. Prime-boost or single shot immunizations in C57BL/6 mice readily induced circulating serum antibodies’ binding to recombinant IAV-NP and the robust activation of IAV-NP-specific CD8+ T cell responses. Moreover, the MVA-NP candidate vaccines protected C57BL/6 mice against lethal respiratory infection with mouse-adapted IAV (A/Puerto Rico/8/1934/H1N1). Thus, further studies are warranted to evaluate the immunogenicity and efficacy of these recombinant MVA-NP vaccines in other IAV challenge models in more detail.

Mucosal vaccines are designed to elicit both a strong systemic and mucosal immune response gaining importance as the next generation of vaccines to combat the respiratory coronavirus disease 2019 (COVID-19). The ability of these vaccines to induce mucosal immune responses in the upper respiratory tract may allow efficient prevention of infection and transmission, which could potentially reduce virus circulation in the population. In addition, they have the advantage that they can be administered by non-medical personnel and without needles. Several preclinical studies in small animal models and non-human primates, but also early phase clinical studies confirmed the capability of mucosal COVID-19 candidate vaccines to induce long-lasting immunity and to provide protection against an infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). In addition to five vaccines which are already approved/licensed, at least 28 mucosal COVID-19 vaccines, based on different vaccine platforms, are currently being tested in clinical trials. However, clinical data for most of the conducted and completed clinical trials are not publicly available yet. In addition, several initiated trials have been terminated or were withdrawn. In this review, we aim to discuss the advantages and disadvantages of mucosal COVID-19 vaccines and to summarize the current status of mucosal COVID-19 vaccines in clinical development, with an emphasis on the most advanced candidates and the key findings observed in preclinical animal models and clinical studies.

Brucellosis, mainly caused by Brucella (B.) melitensis, is associated with a risk of chronification and relapses. Antimicrobial susceptibility testing (AST) standards for B. melitensis are not available, and the agent is not yet listed in the EUCAST breakpoint tables. CLSI recommendations for B. melitensis exist, but they do not fulfill the requirements of the ISO 20776 standard regarding the culture medium and the incubation conditions. Under the third EU Health Programme, laboratories specializing in the diagnostics of highly pathogenic bacteria in their respective countries formed a working group within a Joint Action aiming to develop a suitable method for the AST of B. melitensis. Under the supervision of EUCAST representatives, this working group adapted the CLSI M45 document to the ISO 20776 standard after testing and validation. These adaptations included the comparison of various culture media, culture conditions and AST methods. A Standard Operation Procedure was derived and an interlaboratory validation was performed in order to evaluate the method. The results showed pros and cons for both of the two methods but also indicate that it is not necessary to abandon Mueller–Hinton without additives for the AST of B. melitensis.

The COVID-19 pandemic caused significant human health and economic consequences. Due to the ability of SARS-CoV-2 to spread rapidly and to cause severe disease and mortality in certain population groups, vaccines are essential for controlling the pandemic in the future. Several licensed vaccines have shown improved protection against SARS-CoV-2 after extended-interval prime-boost immunizations in humans. Therefore, in this study, we aimed to compare the immunogenicity of our two Modified Vaccinia virus Ankara (MVA) based COVID-19 candidate vaccines MVA-SARS-2-S and MVA-SARS-2-ST after short- and long-interval prime-boost immunization schedules in mice. We immunized BALB/c mice using 21-day (short-interval) or 56-day (long-interval) prime-boost vaccination protocols and analyzed spike (S)-specific CD8 T cell immunity and humoral immunity. The two schedules induced robust CD8 T cell responses with no significant differences in their magnitude. Furthermore, both candidate vaccines induced comparable levels of total S, and S2-specific IgG binding antibodies. However, MVA-SARS-2-ST consistently elicited higher amounts of S1-, S receptor binding domain (RBD), and SARS-CoV-2 neutralizing antibodies in both vaccination protocols. Overall, we found very comparable immune responses following short- or long-interval immunization. Thus, our results suggest that the chosen time intervals may not be suitable to observe potential differences in antigen-specific immunity when testing different prime-boost intervals with our candidate vaccines in the mouse model. Despite this, our data clearly showed that MVA-SARS-2-ST induced superior humoral immune responses relative to MVA-SARS-2-S after both immunization schedules.