Influenza virus inactivated by artificial ribonucleases as a prospective killed virus vaccine

The inactivation of viral particles with agents causing minimal damage to the structure of surface epitopes is a well-established approach for the production of killed virus vaccines. Here, we describe new agents for the inactivation of influenza virus, artificial ribonucleases (aRNases), which are chemical compounds capable of cleaving RNA molecules. Several aRNases were identified, exhibiting significant virucidal activity against the influenza A virus and causing a minimal effect on the affinity of monoclonal antibodies for the inactivated virus. Using a murine model of the influenza virus infection, a high protective activity of the aRNase-inactivated virus as a vaccine was demonstrated. The results of the experiments demonstrate the efficacy of novel chemical agents in the preparation of vaccines against influenza and, perhaps, against other infections caused by RNA viruses.     

Comparative genome analysis and molecular epidemiology of the reemerging porcine epidemic diarrhea virus strains isolated in Korea

Porcine epidemic diarrhea virus (PEDV), a member of the Coronaviridae family, is an enveloped, positive-sense, single-stranded RNA virus, which causes severe diarrhea and dehydration in suckling pigs. We detected three PEDV strains from ten small intestine samples from piglets with acute diarrhea and we determined the complete genome sequences of the reemerging Korean PEDV field isolates, except for the noncoding regions from both ends. The complete genome sequences of the strains were identical or almost identical (one synonymous single-nucleotide polymorphism (SNP) in the ORF1a/1b genomic sequence). Interestingly, comparative genome analysis of recent Korean PEDV isolates and other strains revealed that the complete genome sequences of recent Korean strains were almost identical (99.9%) to those of the US PEDV strains isolated in 2013. These results suggest that the three reemerging Korean strains are distinct from previous endemic Korean PEDV strains and has been recently introduced into Korea from oversea with high likelihood.     

Sendai virus-based RSV vaccine protects African green monkeys from RSV infection

Respiratory syncytial virus (RSV) is a serious disease of children, responsible for an estimated 160,000 deaths per year worldwide. Despite the ongoing need for global prevention of RSV and decades of research, there remains no licensed vaccine. Sendai virus (SeV) is a mouse parainfluenza virus-type 1 which has been previously shown to confer protection against its human cousin, human parainfluenza virus-type 1 in African green monkeys (AGM). Here is described the study of a RSV vaccine (SeVRSV), produced by reverse genetics technology using SeV as a backbone to carry the full-length gene for RSV F. To test for immunogenicity, efficacy and safety, the vaccine was administered to AGM by intratracheal (i.t.) and intranasal (i.n.) routes. Control animals received the empty SeV vector or PBS. There were no booster immunizations. SeV and SeVRSV were cleared from the URT and LRT of vaccinated animals by day 10. Antibodies with specificities toward SeV and RSV were detected in SeVRSV primed animals as early as day ten after immunizations in both sera and nasal wash samples. One month after immunization all test and control AGM received an i.n. challenge with RSV-A2. SeVRSV-vaccinated animals exhibited reduced RSV in the URT compared to controls, and complete protection against RSV in the LRT. There were no clinically relevant adverse events associated with vaccination either before or after challenge. These data encourage advanced testing of the SeVRSV vaccine candidate in clinical trials for protection against RSV.     

Chinese-like Strain of Porcine Epidemic Diarrhea Virus, Thailand

Since late 2007, several outbreaks of porcine epidemic diarrhea virus (PEDV) infection have emerged in Thailand. Phylogenetic analysis places all Thai PEDV isolates during the outbreaks in the same clade as the Chinese strain JS-2004-2. This new genotype PEDV is prevailing and currently causing sporadic outbreaks in Thailand.     

Sendai virus-based RSV vaccine protects against RSV challenge in an in vivo maternal antibody model

Respiratory syncytial virus (RSV) is the cause of significant morbidity and mortality among infants, and despite decades of research there remains no licensed vaccine. SeVRSV is a Sendai virus (SeV)-based live intranasal vaccine that expresses the full length RSV fusion (F) gene. SeV is the murine counterpart of human parainfluenza virus type 1. Given that the target population of SeVRSV is young infants, we questioned whether maternal antibodies typical of this age group would inhibit SeVRSV vaccine efficacy. After measuring SeV- and RSV-specific serum neutralizing antibody titers in human infants, we matched these defined titers in cotton rats by the passive transfer of polyclonal or monoclonal antibody products. Animals were then vaccinated with SeVRSV followed by a 3 month rest period to allow passively transferred antibodies to wane. Animals were finally challenged with RSV to measure the de novo vaccine-induced immune responses. Despite the presence of passively-transferred serum neutralizing antibodies at the time of vaccination, SeVRSV induced immune responses that were protective against RSV challenge. The data encourage advancement of SeVRSV as a candidate vaccine for the protection of children from morbidity and mortality caused by RSV.     

Enhanced protective immunity to CSFV E2 subunit vaccine by using IFN-γ as immunoadjuvant in weaning piglets

The glycoprotein E2 of classical swine fever virus (CSFV) is a major immunogenic protein that induces neutralizing antibodies and protective immunity. Thus, E2 is a suitable target antigen for the development of genetically engineered CSFV vaccines. However, these vaccines cannot generate complete protective immunity in their hosts, thereby limiting the scope of applications under field conditions. IFN-γ is an immune adjuvant that has been shown to enhance antigen immune response in various experimental models. In this study, porcine IFN-γ was used to improve the immunogenicity of the CSFV E2 subunit vaccine in pigs. Pigs were immunized with E2 subunit vaccine alone or in combination with IFN-γ. Results demonstrated that porcine IFN-γ did not enhance the CSFV-specific antibody and neutralizing antibody titers compared with the E2 subunit vaccine alone. However, co-administration of the E2 and IFN-γ subunit vaccines significantly enhanced the CSFV-specific IFN-γ expression. These findings indicated that porcine IFN-γ can increase cellular immune responses to E2 protein in pigs. Furthermore, co-immunization with E2 + IFN-γ subunit vaccine and C-strain conferred complete protection against CSFV. In contrast, E2 subunit vaccines provided incomplete protection in pigs. These results indicated that using IFN-γ as an adjuvant with CSFV E2 subunit vaccines can enhance the specific protective immune response. Therefore, E2 + IFN-γ subunit vaccine is a promising marker vaccine candidate for the control and eradication of CSF.     

A novel subunit vaccine co-expressing GM-CSF and PCV2b Cap protein enhances protective immunity against porcine circovirus type 2 in piglets

Porcine circovirus type 2 (PCV2) causes porcine circovirus-associated disease. Capsid (Cap) protein of PCV2 is the principal immunogenic protein that induces neutralizing antibodies and protective immunity. GM-CSF is an immune adjuvant that enhances responses to vaccines. In this study, recombinant baculoviruses Ac-Cap and Ac-Cap-GM-CSF expressing the Cap protein alone and co-expressing the Cap protein and porcine GM-CSF, respectively, were constructed successfully. The target proteins were analyzed by western blotting and IFA. Further, these proteins were confirmed by electron microscopy, which showed that Cap proteins could self-assemble into virus-like particles having diameters of 17–25 nm. Animal experiments showed that pigs immunized with Cap-GM-CSF subunit vaccine showed significantly higher levels of PCV2-specific antibodies and neutralizing antibodies than pigs immunized with the Cap subunit vaccine and a commercial vaccine (Ingelvac CircoFLEX; P < 0.05). After PCV2 wild strain challenged, Pigs receiving the Cap-GM-CSF subunit vaccine showed significantly higher average daily weight gain after wild-type PCV2 challenge than pigs receiving the other three vaccines (P < 0.05). None of PCV2 DNA was detected in all immunized animals, except control animals immunized with phosphate-buffered saline. These results indicated that GM-CSF was a powerful immunoadjuvant for PCV2 subunit vaccines because it enhanced humoral immune response and improved immune protection against PCV2 infection in pigs. Thus, the novel Cap-GM-CSF subunit vaccine has the potential to be used as an effective and safe vaccine candidate against PCV2 infection.     

Porcine Epidemic Diarrhea Virus and Discovery of a Recombinant Swine Enteric Coronavirus,Italy

Porcine epidemic diarrhea virus (PEDV) has been detected sporadically in Italy since the 1990s. We report the phylogenetic relationship of swine enteric coronaviruses collected in Italy during 2007–2014 and identify a drastic shift in PEDV strain variability and a new swine enteric coronavirus generated by recombination of transmissible gastroenteritis virus and PEDV.     

Novel Porcine Epidemic Diarrhea Virus Variant with Large Genomic Deletion,South Korea

Since 1992, porcine epidemic diarrhea virus (PEDV) has been one of the most common porcine diarrhea–associated viruses in South Korea. We conducted a large-scale investigation of the incidence of PEDV in pigs with diarrhea in South Korea and consequently identified and characterized a novel PEDV variant with a large genomic deletion.     

Making Better Influenza Virus Vaccines?

Killed and live influenza virus vaccines are effective in preventing and curbing the spread of disease, but new technologies such as reverse genetics could be used to improve them and to shorten the lengthy process of preparing vaccine seed viruses. By taking advantage of these new technologies, we could develop live vaccines that would be safe, cross-protective against variant strains, and require less virus per dose than conventional vaccines. Furthermore, pandemic vaccines against highly virulent strains such as the H5N1 virus can only be generated by reverse genetics techniques. Other technologic breakthroughs should result in effective adjuvants for use with killed and live vaccines, increasing the number of available doses. Finally, universal influenza virus vaccines seem to be within reach. These new strategies will be successful if they are supported by regulatory agencies and if a robust market for influenza virus vaccines against inter-pandemic and pandemic threats is made and sustained.     

Presentation of peptides from Bacillus anthracis protective antigen on Tobacco Mosaic Virus as an epitope targeted anthrax vaccine

The current anthrax vaccine requires improvements for rapidly invoking longer-lasting neutralizing antibody responses with fewer doses from a well-defined formulation. Designing antigens that target neutralizing antibody epitopes of anthrax protective antigen, a component of anthrax toxin, may offer a solution for achieving a vaccine that can induce strong and long lasting antibody responses with fewer boosters. Here we report implementation of a strategy for developing epitope focused virus nanoparticle vaccines against anthrax by using immunogenic virus particles to present peptides derived from anthrax toxin previously identified in (1) neutralizing antibody epitope mapping studies, (2) toxin crystal structure analyses to identify functional regions, and (3) toxin mutational analyses. We successfully expressed two of three peptide epitopes from anthrax toxin that, in previous reports, bound antibodies that were partially neutralizing against toxin activity, discovered cross-reactivity between vaccine constructs and toxin specific antibodies raised in goats against native toxin and showed that antibodies induced by our vaccine constructs also cross-react with native toxin. While protection against intoxication in cellular and animal studies were not as effective as in previous studies, partial toxin neutralization was observed in animals, demonstrating the feasibility of using plant-virus nanoparticles as a platform for epitope defined anthrax vaccines.