Motivationsanalyse von Touristen in Wien

Studien, die sich mit dem Thema Städtetourismus insbesondere in Verbindung mit Motivation auseinandersetzen, sind äußerst selten. Die vorliegende Arbeit untersucht daher Zusammenhänge zwischen Motiven (Intellektuelle-, Soziale-, Kompetenz-Macht- und Stimulusvermeidungsmotiven) und Urlaubsaktivitäten von Touristen in Wien. Zur Erfassung der Motive wird die „Leisure Motivation Scale“ von Beard und Ragheb (1983) in adaptierter Form eingesetzt. Neben den Motiven werden soziodemografische und reisebezogene Variablen auf ihre Bedeutung für Urlaubsaktivitäten überprüft. Aus marktwirtschaftlichen Interessen werden die Urlauber anhand einer Clusteranalyse aufgrund ihre Urlaubsaktivitäten in drei Typen (Sightseeing-, Sport und Fun-, Intellektueller Typ) eingeteilt, wobei sich mittels Diskriminanzanalyse zeigt, dass 64,6 % der Urlauber aufgrund weniger Variablen (Sozial Motiv, gesamte Aufenthaltsdauer, Alter) zu den Urlaubstypen richtig zugeordnet werden können

Monoclonal antibodies targeting the influenza virus N6 neuraminidase

Influenza A viruses are a diverse species that include 16 true hemagglutinin (HA) subtypes and 9 true neuraminidase (NA) subtypes. While the antigenicity of many HA subtypes is reasonably well studied, less is known about NA antigenicity, especially when it comes to non-human subtypes that only circulate in animal reservoirs. The N6 subtype NAs are mostly found in viruses infecting birds. However, they have also been identified in viruses that infect mammals, such as swine and seals. More recently, highly pathogenic H5N6 subtype viruses have caused rare infections and mortality in humans. Here, we generated murine mAbs to the N6 NA, characterized their breadth and antiviral properties in vitro and in vivo and mapped their epitopes by generating escape mutant viruses. We found that the antibodies had broad reactivity across the American and Eurasian N6 lineages, but relatively little binding to the H5N6 NA. Several of the antibodies exhibited strong NA inhibition activity and some also showed activity in the antibody dependent cellular cytotoxicity reporter assay and neutralization assay. In addition, we generated escape mutant viruses for six monoclonal antibodies and found mutations on the lateral ridge of the NA. Lastly, we observed variable protection in H4N6 mouse challenge models when the antibodies were given prophylactically

Increased efficacy of influenza virus vaccine candidate through display of recombinant neuraminidase on virus like particles

Vaccination against influenza virus can reduce the risk of influenza by 40% to 60%, they rely on the production of neutralizing antibodies specific to influenza hemagglutinin (HA) ignoring the neuraminidase (NA) as an important surface target. Vaccination with standardized NA concentration may offer broader and longer-lasting protection against influenza infection. In this regard, we aimed to compare the potency of a NA displayed on the surface of a VLP with a soluble NA. The baculovirus expression system (BEVS) and the novel virus-free Tnms42 insect cell line were used to express N2 NA on gag-based VLPs. To produce VLP immunogens with high levels of purity and concentration, a two-step chromatography purification process combined with ultracentrifugation was used. In a prime/boost vaccination scheme, mice vaccinated with 1 µg of the N2-VLPs were protected from mortality, while mice receiving the same dose of unadjuvanted NA in soluble form succumbed to the lethal infection. Moreover, NA inhibition assays and NA-ELISAs of pre-boost and pre-challenge sera confirm that the VLP preparation induced higher levels of NA-specific antibodies outperforming the soluble unadjuvanted NA

Heterologous prime-boost vaccination with H3N2 influenza viruses of swine favors cross-clade antibody responses and protection

The emergence of multiple novel lineages of H1 and H3 influenza A viruses in swine has confounded control by inactivated vaccines. Because of substantial genetic and geographic heterogeneity among circulating swine influenza viruses, one vaccine strain per subtype cannot be efficacious against all of the current lineages. We have performed vaccination-challenge studies in pigs to examine whether priming and booster vaccinations with antigenically distinct H3N2 swine influenza viruses could broaden antibody responses and protection. We prepared monovalent whole inactivated, adjuvanted vaccines based on a European and a North American H3N2 swine influenza virus, which showed 81.5% aa homology in the HA1 region of the hemagglutinin and 83.4% in the neuraminidase. Our data show that (i) Priming with European and boosting with North American H3N2 swine influenza virus induces antibodies and protection against both vaccine strains, unlike prime-boost vaccination with a single virus or a single administration of bivalent vaccine. (ii) The heterologous prime-boost vaccination enhances hemagglutination inhibiting, virus neutralizing and neuraminidase inhibiting antibody responses against H3N2 viruses that are antigenically distinct from both vaccine strains. Antibody titers to the most divergent viruses were higher than after two administrations of bivalent vaccine. (iii) However, it does not induce antibodies to the conserved hemagglutinin stalk or to other hemagglutinin subtypes. We conclude that heterologous prime-boost vaccination might broaden protection to H3N2 swine influenza viruses and reduce the total amount of vaccine needed. This strategy holds potential for vaccination against influenza viruses from both humans and swine and for a better control of (reverse) zoonotic transmission of influenza viruses

Development of a universal group 2 influenza virus vaccine using chimeric hemagglutinin constructs

The stalk domain of the hemagglutinin (HA) has become the prime target for universal influenza virus vaccine development in the last few years. Unlike the HA head domain, the immunosubdominant stalk domain is conserved to a higher level within each influenza virus HA group. Sequential vaccination with chimeric HA (cHA) vaccine constructs consisting of the same HA stalk and exotic head domains has proven to re-direct the immune response towards the stalk domain. This vaccination concept provides the basis for the development of more broadly cross-protective vaccines that are less affected by antigenic drift and shift, one of the main drawbacks of currently marketed influenza vaccines. Most influenza virus vaccines are licensed as inactivated split vaccines. They are manufactured based on HA content with little to no information and standardization of neuraminidase (NA) content. Virus inactivation is generally performed with alkylating agents such as formalin (FA) or β-propiolactone (βPL), rendering the virus unable to infect or replicate. Though safe, whole inactivated virus vaccines can be highly reactogenic. Virus splitting with detergents like sodium deoxycholate (SDCO) and Triton X-100 (TX-100), which dissociate the virus into smaller parts while maintaining a good immunogenicity profile, are typically employed. To date, there are several studies assessing the effect of a variety of inactivating and splitting agents on influenza viruses, but little is known about the impact of combining these agents on HA stalk conformation and NA activity. Please click Download on the upper right corner to see the full abstract

Production and characterization of HER2-displaying budded virus-like particles and their potential as cancer vaccines

Cancer biomarkers are often glycosylated membrane receptor proteins present on the cellular surface. In order to utilize such receptor proteins in designing specific and sensitive diagnostic tools or as immunogens for vaccination based treatments, they need to be expressed in their native conformation. However, membrane receptor proteins are notoriously difficult to produce due to their hydrophobic nature and complex structure. The human epidermal growth factor receptor 2 (HER2) is known to be up regulated in a number of cancers including breast cancer, lung cancer, gastric cancer and glioblastoma multiform and was therefore chosen as tumor antigen in our studies. Here we used the baculovirus-insect cell expression vector system (BEVS) to produce budded virus-like particles (VLPs) serving as a display platform for the antigen. VLPs displaying HER2 were produced in Spodoptera frugiperda (Sf9) insect cells and were purified by sucrose gradient ultracentrifugation. The number of secreted particles was quantified by nanoparticle tracking analysis. To confirm the presence and functionality of displayed HER2, VLPs were labeled with gold-conjugated antibodies, were analyzed by transmission electron microscopy and the ability to present native epitopes was tested through enzyme-linked immunosorbent assay (ELISA). Trastuzumab, an anti-HER2 antibody, showed significant binding to antigen displaying VLPs, which demonstrates the potential of this platform to display cell surface biomarkers in their authentic conformation. In the second part of the study, the efficacy of the aforementioned characterized VLPs as a cancer vaccine was investigated. BALB/c mice were injected intramuscularly with control VLPS and HER2-displaying VLPs in combination with two different adjuvants in a prime-boost regimen. As verified by ELISA, HER2-displaying VLP vaccines induced strong antibody responses when tested against recombinant HER2, with variability observed amongst the different adjuvant groups. For further characterization the antibody-dependent cell-mediated cytotoxicity (ADCC) potential of the induced antibodies will be investigated and vaccinated mice will be challenged with HER2 expressing tumors to test the potential of antigen-displaying VLPs as a cancer vaccine. Overall, using our strategy, many other membrane proteins including tumor antigens, immune cell markers and immune receptors could be expressed. These tools could further be instrumental in cancer vaccine design and diagnostics, as well as antibody selection and engineering

Novel purification strategies for Influenza neuraminidase-VLPs

Conventional Influenza virus vaccines reduce the risk of flu illness by between 40% and 60% among the overall population. They are all standardized based on the influenza hemagglutinin (HA) content and focus on the induction of HA-specific neutralizing antibodies. However, in contrast to natural infection, seasonal vaccination fails to induce an efficient immune response to the Influenza virus neuraminidase (NA), the second most abundant viral glycoprotein. A potential solution to this problem would be to supplement current vaccines with a correctly folded, tetrameric recombinant NA protein, or alternatively, with bio-nanoparticles that display the tetrameric NA on an outer membrane. Vaccines that incorporate standardized amounts of NA might lead to broader and longer-lasting protection against influenza infection. Please click Download on the upper right corner to see the full abstract

Persistently high antibody responses after AS03-adjuvanted H1N1pdm09 vaccine: Dissecting the HA specific antibody response

Current influenza vaccines have a suboptimal effectiveness. The introduction of a novel A/H1N1 influenza virus in 2009 (H1N1pdm09) provided a unique opportunity to study the humoral response to the AS03-adjuvanted H1N1pdm09 vaccine and repeated annual vaccination with the homologous virus in subsequent influenza seasons. Thirty-two HCWs immunized with the AS03-adjuvanted H1N1pdm09 vaccine in 2009 were divided into four groups based on the longevity of their antibody responses (persistently high or transient), and whether they were repeatedly annually vaccinated in the subsequent four influenza seasons or not. Serological assays were utilized to measure the quantity, quality and functionality of antibodies targeting the major surface glycoprotein hemagglutinin (HA). Persistent high responders (hemagglutination inhibition (HI) titre ≥ 80 at 12 months after H1N1pdm09 vaccination) had protective levels of HI antibodies throughout the study period. In addition, the quality and functionality of these antibodies were greater than the individuals who had a transient antibody response to the pandemic vaccine (HI titre < 40 at 12 months after H1N1pdm09 vaccination). All groups had similar levels of antibodies towards the conserved HA stalk domain. The level of HA head-specific antibodies gradually increased over time with annual vaccination in the transient responders. The AS03-adjuvanted H1N1pdm09 vaccine elicited a robust humoral response that persisted up to 5 years in some individuals. Seasonal annual vaccination boosted the HA-antibodies over time in individuals with a transient response to the pandemic H1N1pdm09 vaccine.publishedVersio

Bioprocess development for universal influenza vaccines based on inactivated split chimeric and mosaic hemagglutinin viruses

Seasonal influenza viruses account for 1 billion infections worldwide every year, including 3–5 million cases of severe illness and up to 650,000 deaths. The effectiveness of current influenza virus vaccines is variable and relies on the immunodominant hemagglutinin (HA) and to a lesser extent on the neuraminidase (NA), the viral surface glycoproteins. Efficient vaccines that refocus the immune response to conserved epitopes on the HA are needed to tackle infections by influenza virus variants. Sequential vaccination with chimeric HA (cHA) and mosaic HA (mHA) constructs has proven to induce immune responses to the HA stalk domain and conserved epitopes on the HA head. In this study, we developed a bioprocess to manufacture cHA and mHA inactivated split vaccines and a method to quantify HA with a prefusion stalk based on a sandwich enzyme-linked immunosorbent assay. Virus inactivation with beta-propiolactone (βPL) and splitting with Triton X-100 yielded the highest amount of prefusion HA and enzymatically active NA. In addition, the quantity of residual Triton X-100 and ovalbumin (OVA) was reduced to very low levels in the final vaccine preparations. The bioprocess shown here provides the basis to manufacture inactivated split cHA and mHA vaccines for pre-clinical research and future clinical trials in humans, and can also be applied to produce vaccines based on other influenza viruses

Prolonging the delivery of influenza virus vaccine improves the quantity and quality of the induced immune responses in mice

IntroductionInfluenza vaccines play a vital role in protecting individuals from influenza virus infection and severe illness. However, current influenza vaccines have suboptimal efficacy, which is further reduced in cases where the vaccine strains do not match the circulating strains. One strategy to enhance the efficacy of influenza vaccines is by extended antigen delivery, thereby mimicking the antigen kinetics of a natural infection. Prolonging antigen availability was shown to quantitatively enhance influenza virus-specific immune responses but how it affects the quality of the induced immune response is unknown. Therefore, the current study aimed to investigate whether prolongation of the delivery of influenza vaccine improves the quality of the induced immune responses over that induced by prime-boost immunization.MethodsMice were given daily doses of whole inactivated influenza virus vaccine for periods of 14, 21, or 28 days; the control group received prime-boost immunization with a 28 days interval.ResultsOur data show that the highest levels of cellular and humoral immune responses were induced by 28 days of extended antigen delivery, followed by 21, and 14 days of delivery, and prime-boost immunization. Moreover, prolonging vaccine delivery also improved the quality of the induced antibody response, as indicated by higher level of high avidity antibodies, a balanced IgG subclass profile, and a higher level of cross-reactive antibodies.ConclusionsOur findings contribute to a better understanding of the immune response to influenza vaccination and have important implications for the design and development of future slow-release influenza vaccines