An inactivated Vero cell-grown Japanese encephalitis vaccine formulated with Advax, a novel inulin-based adjuvant, induces protective neutralizing antibody against homologous and heterologous flaviviruses

Advax is a polysaccharide-based adjuvant that potently stimulates vaccine immunogenicity without the increased reactogenicity seen with other adjuvants. This study investigated the immunogenicity of a novel Advax-adjuvanted Vero cell culture candidate vaccine against Japanese encephalitis virus (JEV) in mice and horses. The results showed that, in mice, a two-immunization, low-dose (50 ng JEV antigen) regimen with adjuvanted vaccine produced solid neutralizing immunity comparable to that elicited with live ChimeriVax-JE immunization and superior to that elicited with tenfold higher doses of a traditional non-adjuvanted JEV vaccine (JE-VAX; Biken Institute) or a newly approved alum-adjuvanted vaccine (Jespect; Novartis). Mice vaccinated with the Advax-adjuvanted, but not the unadjuvanted vaccine, were protected against live JEV challenge. Equine immunizations against JEV with Advax-formulated vaccine similarly showed enhanced vaccine immunogenicity, confirming that the adjuvant effects of Advax are not restricted to rodent models. Advax-adjuvanted JEV vaccine elicited a balanced T-helper 1 (Th1)/Th2 immune response against JEV with protective levels of cross-neutralizing antibody against other viruses belonging to the JEV serocomplex, including Murray Valley encephalitis virus (MVEV). The adjuvanted JEV vaccine was well tolerated with minimal reactogenicity and no systemic toxicity in immunized animals. The cessation of manufacture of traditional mouse brain-derived unadjuvanted JEV vaccine in Japan has resulted in a JEV vaccine shortage internationally. There is also an ongoing lack of human vaccines against other JEV serocomplex flaviviruses, such as MVEV, making this adjuvanted, cell culture-grown JEV vaccine a promising candidate to address both needs with one vaccine

West Nile virus and its emergence in the United States of America

Zoonotic West Nile virus (WNV) circulates in natural transmission cycles involving certain mosquitoes and birds, horses, humans, and a range of other vertebrates are incidental hosts. Clinical infections in humans can range in severity from uncomplicated WNV fever to fatal meningoencephalitis. Since its introduction to the Western Hemisphere in 1999, WNV had spread across North America, Central and South America and the Caribbean, although the vast majority of severe human cases have occurred in the United States of America (USA) and Canada. By 2002–2003, the WNV outbreaks have involved thousands of patients causing severe neurologic disease (meningoencephalitis and poliomyelitis-like syndrome) and hundreds of associated fatalities in USA. The purpose of this review is to present recent information on the epidemiology and pathogenicity of WNV since its emergence in North America

MHC class II-alpha chain knockout mice support increased viral replication that is independent of their lack of MHC class II cell surface expression and associated immune function deficiencies

MHCII molecules are heterodimeric cell surface proteins composed of an α and β chain. These molecules are almost exclusively expressed on thymic epithelium and antigen presenting cells (APCs) and play a central role in the development and function of CD4 T cells. Various MHC-II knockout mice have been generated including MHC-IIAα-/- (I-Aα-/-), MHC-IIAβ-/- (I-β-/-) and the double knockout (I-Aαxβ-/-). Here we report a very striking observation, namely that alphaviruses including the avirulent strain of Semliki Forest virus (aSFV), which causes asymptomatic infection in wild-type C57BL6/J (B6) mice, causes a very acute and lethal infection in I-Aα-/-, but not in I-β-/- or I-Aαxβ-/-, mice. This susceptibility to aSFV is associated with high virus titres in muscle, spleen, liver, and brain compared to B6 mice. In addition, I-Aα-/- mice show intact IFN-I responses in terms of IFN-I serum levels and IFN-I receptor expression and function. Radiation bone marrow chimeras of B6 mice reconstituted with I-Aα-/- bone marrow expressed B6 phenotype, whereas radiation chimeras of I-Aα-/- mice reconstituted with B6 bone marrow expressed the phenotype of high viral susceptibility. Virus replication experiments both in vivo and in vitro showed enhanced virus growth in tissues and cell cultures derived form I-Aα-/- compared to B6 mice. This enhanced virus replication is evident for other alpha-, flavi- and poxviruses and may be of great benefit to producers of viral vaccines. In conclusion, I-Aα-/- mice exhibit a striking susceptibility to virus infections independent of their defective MHC-II expression. Detailed genetic analysis will be carried out to characterise the underlining genetic defects responsible for the observed phenomenon.Mohammed Alsharifi, Aulikki Koskinen, Danushka K. Wijesundara, Jayaram Bettadapura, Arno Müllbache

Lack of both Fas Ligand and Perforin Protects from Flavivirus-Mediated Encephalitis in Mice

The mechanism by which encephalitic flaviviruses enter the brain to inflict a life-threatening encephalomyelitis in a small percentage of infected individuals is obscure. We investigated this issue in a mouse model for flavivirus encephalitis in which the virus was administered to 6-week-old animals by the intravenous route, analogous to the portal of entry in natural infections, using a virus dose in the range experienced following the bite of an infectious mosquito. In this model, infection with 0.1 to 10(5) PFU of virus gave mortality in ∼50% of animals despite low or undetectable virus growth in extraneural tissues. We show that the cytolytic effector functions play a crucial role in invasion of the encephalitic flavivirus into the brain. Mice deficient in either the granule exocytosis- or Fas-mediated pathway of cytotoxicity showed delayed and reduced mortality. Mice deficient in both cytotoxic effector functions were resistant to a low-dose peripheral infection with the neurotropic virus

Using Citizen Science to Scout Honey Bee Colonies That Naturally Survive Varroa destructor Infestations

Simple Summary Citizen Science is a valuable resource that can substantially contribute to the conservation of biodiversity. However, its use in honey bee research has remained minimal. The Survivors Task Force of the COLOSS association created and promoted an online surveying tool with the aim of identifying potential cases of Western honey bee, Apis mellifera, populations that are surviving infestations with ectoparasitic mites Varroa destructor without control measures by beekeepers. The reports suggest that there could be twice as many naturally surviving colonies worldwide than are currently known. The survey also shows that citizens can be readily engaged through social media, personal networks, and promotional campaigns to gather valuable and previously inaccessible data. These reports of surviving honey bee colonies will now be validated through the new initiative Honey Bee Watch, a global and multi-year Citizen Science project founded to connect citizens, beekeepers, and scientists. This will enable to increase scientific knowledge, mitigate honey bee colony losses, and develop education and conservation campaigns. Citizen Science contributes significantly to the conservation of biodiversity, but its application to honey bee research has remained minimal. Even though certain European honey bee (Apis mellifera) populations are known to naturally survive Varroa destructor infestations, it is unclear how widespread or common such populations are. Such colonies are highly valuable for investigating the mechanisms enabling colony survival, as well as for tracking the conservation status of free-living honey bees. Here, we use targeted Citizen Science to identify potentially new cases of managed or free-living A. mellifera populations that survive V. destructor without mite control strategies. In 2018, a survey containing 20 questions was developed, translated into 13 languages, and promoted at beekeeping conferences and online. After three years, 305 reports were collected from 28 countries: 241 from managed colonies and 64 from free-living colonies. The collected data suggest that there could be twice as many naturally surviving colonies worldwide than are currently known. Further, online and personal promotion seem to be key for successful recruitment of participants. Although the survivor status of these colonies still needs to be confirmed, the volume of reports and responses already illustrate how effectively Citizen Science can contribute to bee research by massively increasing generated data, broadening opportunities for comparative research, and fostering collaboration between scientists, beekeepers, and citizens. The success of this survey spurred the development of a more advanced Citizen Science platform, Honey Bee Watch, that will enable a more accurate reporting, confirmation, and monitoring of surviving colonies, and strengthen the ties between science, stakeholders, and citizens to foster the protection of both free-living and managed honey bees

In situ reactions of monoclonal antibodies with a viable mutant of Murray Valley encephalitis virus reveal an absence of dimeric NS1 protein

Studies on the NS1 protein of flaviviruses have concluded that formation of a stable homodimer is required for virus replication. However, previous work has reported that substitution of a conserved proline by leucine at residue 250 in NS1 of Kunjin virus (KUNV) eliminated dimerization, but allowed virus replication to continue. To assess whether this substitution has similar effects on other flaviviruses, it was introduced into an infectious clone of Murray Valley encephalitis virus (MVEV). Consistent with studies of KUNV, the mutant virus (MVEVNS1-250Leu) produced high levels of monomeric NS1 and the NS1 homodimer could not be detected. In contrast, wild-type MVEV cultures contained predominantly dimeric NS1. Retarded virus growth in Vero cells and loss of neuroinvasiveness for weanling mice revealed further similarities between MVEVNS1-250Leu and the corresponding KUNV mutant. To confirm that the lack of detection of dimeric NS1 in mutant virus samples was not due to denaturation of unstable dimers during Western blotting, a mAb (2E3) specific for the MVEV NS1 homodimer was produced. When NS1 protein was fixed in situ in mammalian and arthropod cells infected with wild-type or mutant virus, 2E3 reacted strongly with the former, but not the latter These results confirmed that Pro(250) in NS1 is important for dimerization and that substitution of this residue by leucine represents a conserved marker of attenuation for viruses of the Japanese encephalitis virus serocomplex. The inability to detect dimeric NS1 in supernatant or cell monolayers of cultures productively infected with mutant virus also suggests that dimerization of the protein may not be essential for virus replication

A flavivirus signal peptide balances the catalytic activity of two proteases and thereby facilitates virus morphogenesis

10.1016/j.virol.2010.02.008Virology401180-8