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African swine fever virus (ASFV) causes a highly contagious hemorrhagic disease that affects domestic pig and Eurasian wild boar populations. To date, no safe and efficacious treatment or vaccine against ASF is available. Nevertheless, there are several reports of protection elicited by experimental vaccines based on live attenuated ASFV and some levels of protection and reduced viremia in other approaches such as DNA, adenovirus, baculovirus, and vaccinia-based vaccines. Current ASF subunit vaccine research focuses mainly on delivering protective antigens and antigen discovery within the ASFV genome. However, due to the complex nature of ASFV, expression vectors need to be optimized to improve their immunogenicity. Therefore, in the present study, we constructed several recombinant MVA vectors to evaluate the efficiency of different promoters and secretory signal sequences in the expression and immunogenicity of the p30 protein from ASFV. Overall, the natural poxvirus PrMVA13.5L promoter induced high levels of both p30 mRNA and specific anti-p30 antibodies in mice. In contrast, the synthetic PrS5E promoter and the S E/L promoter linked to a secretory signal showed lower mRNA levels and antibodies. These findings indicate that promoter selection may be as crucial as the antigen used to develop ASFV subunit vaccines using MVA as the delivery vector. [ABSTRACT FROM AUTHOR]

The article discusses a research study on characterizing the vectors of recombinant raccoonpox vaccine in chickens. A discussion on previous research studies on poxviruses is detailed. Researchers studied the replicated raccoonpox virus (RCN) in chick embryo fibroblasts (CEF) using the highly pathogenic avian influenza virus (H5N1) hemagglutinin (HA) as model antigen while the reporter antigen used was luciferase (luc). Results of the study indicated that RCN has the potential to be used as vaccine vector for avian influenza and poultry pathogens.

In the present study, we evaluated both the immunogenicity and safety of recombinant raccoon poxvirus (RCN) as a mucosal vaccine vector for domestic cats. RCN is an orthopoxvirus that was isolated from healthy raccoons and has been used experimentally as a vaccine vector for rabies and other antigens in a variety of species, including raccoons, skunks, foxes, bobcats, rabbits, domestic cats, piglets, sheep and non-human primates. We evaluated the antibody response induced by a recombinant RCN vaccine expressing the rabies-G glycoprotein (RCN/rabies-G) administered to cats by the oral (PO), intranasal (IN), conjunctival (CO) or intranasal/conjunctival (IN/CO) route (dose: 10 8 plaque forming units or PFU). The IN route, either alone or combined with the CO route, induced the highest rabies virus neutralizing antibody (RVNA) titers. The RVNA titers remained high when measured at six months post-vaccination, demonstrating that the recombinant vaccine administered via these routes is very efficient at inducing long-lasting immunity. A dose-response was observed following IN vaccination in cats. Doses of 10 7 PFU induced strong antibody responses in 4 of 5 animals [geometric mean titer: 3.2 (log 10 )]. None of the animals vaccinated with 10 4 PFU developed detectable RVNA titers. In this study, RCN/rabies-G viral shedding was below detectable levels. Nasal, oral and fecal swabs collected from these cats were negative for RCN by both virus isolation and by nested-PCR. In addition, no horizontal transmission of the virus could be detected. Gang-housed sentinel animals for each group did not develop detectable anti-RVNA or -RCN antibodies. To study tissue tropism of recombinant raccoon poxvirus vaccines, a RCN that can express the lacZ gene (RCN/ lacZ ) was constructed. Expression of β-galactosidase (β-gal) was validated in vitro and in mice in vivo . Cats were vaccinated IN with 10 7 PFU of RCN/ lacZ . No histopathological lesions were detected in any of the tissues collected from these cats at 1, 4, 7 and 15 days post-vaccination. In addition, no virus or β-gal expression was detected in any of these tissues. Controls demonstrated that virus could be reisolated from nasal swabs immediately after administration of 10 8 PFU to cats. These results suggest that recombinant RCN vaccines undergo limited replication after intranasal administration in cats that is sufficient to elicit strong, long-lasting systemic antibody responses. [ABSTRACT FROM AUTHOR]

Using a raccoon poxvirus (RCN) expression system, we have developed new recombinant vaccines that can protect mice against lethal plague infection. We tested the effects of a translation enhancer (EMCV-IRES) in combination with a secretory (tPA) signal or secretory (tPA) and membrane anchoring (CHV-gG) signals on in vitro antigen expression of F1 antigen in tissue culture and the induction of antibody responses and protection against Yersinia pestis challenge in mice. The RCN vector successfully expressed the F1 protein of Y. pestis in vitro. In addition, the level of expression was increased by the insertion of the EMCV-IRES and combinations of this and the secretory signal or secretory and anchoring signals. These recombinant viruses generated protective immune responses that resulted in survival of 80% of vaccinated mice upon challenge with Y. pestis. Of the RCN-based vaccines we tested, the RCN-IRES-tPA-YpF1 recombinant construct was the most efficacious. Mice vaccinated with this construct withstood challenge with as many as 1.5 million colony forming units of Y. pestis (7.7×104 LD50). Interestingly, vaccination with F1 fused to the anchoring signal (RCN-IRES-tPA-YpF1-gG) elicited significant anti-F1 antibody titers, but failed to protect mice from plague challenge. Our studies demonstrate, in vitro and in vivo, the potential importance of the EMCV-IRES and secretory signals in vaccine design. These molecular tools provide a new approach for improving the efficacy of vaccines. In addition, these novel recombinant vaccines could have human, veterinary, and wildlife applications in the prevention of plague. [Copyright &y& Elsevier]

We previously generated recombinant poxviruses expressing influenza antigens and studied their efficacy as potential highly pathogenic avian influenza (HPAI) vaccines in mice. While both modified vaccinia Ankara (MVA) and raccoon poxvirus (RCN) expressing hemagglutinin (HA) provided strong protection when administered by parenteral routes, only RCN-neuraminidase (NA) showed promise as a mucosal vaccine. In the present study we evaluated the efficacy of RCN-NA constructs by both intradermal (ID) and intranasal (IN) routes. Surprisingly, while RCN-NA completely protected mice when administered by the IN route, it failed to protect mice when administered by the ID route. After challenge, significantly less virus induced pathology was observed in the lungs of mice vaccinated with RCN-NA by the IN route as compared to the ID route. Furthermore, IN administration of RCN-NA elicited neutralizing antibodies detected in bronchoalveolar lavage (BAL) samples. We also determined the role of cellular immune responses in protection elicited by RCN-NA by depleting CD4 and CD8 T cells prior to challenge. Finally, we demonstrated for the first time that antibodies against NA can block viral entry in addition to viral spread in vitro . These studies demonstrate the importance of mucosal administration of RCN viral vectors for eliciting protective immune responses against the NA antigen. [ABSTRACT FROM AUTHOR]

Abstract: We previously demonstrated that recombinant raccoonpox (RCN) virus could serve as a vector for an influenza vaccine. RCN constructs expressing the hemagglutinin (HA) from H5N1 viruses were immunogenic in chickens. In the current study, we generated several recombinant RCN constructs expressing influenza (H5N1) antigens and a molecular adjuvant (Heat-Labile enterotoxin B from E. coli: RCN-LTB), demonstrated their expression in vitro, and evaluated their ability to protect mice against H5N1 virus challenge. RCN-HA provided strong protection when administered intradermally (ID), but not intranasally (IN). Conversely, the RCN-neuraminidase (NA) construct was highly efficacious by the IN route and elicited high titers of neutralizing antibodies in mice. Vaccination by combined ID (RCN-HA) and IN (RCN-NA) routes offered mice the best protection against an IN challenge with heterologous H5N1 virus. However, protection was reduced when the different RCN constructs were pre-mixed, perhaps due to reduced expression of antigen. [Copyright &y& Elsevier]

Abstract: In previous studies, we demonstrated protection against plague in mice and prairie dogs using a raccoon pox (RCN) virus-vectored vaccine that expressed the F1 capsular antigen of Yersinia pestis. In order to improve vaccine efficacy, we have now constructed additional RCN-plague vaccines containing two different forms of the lcrV (V) gene, including full-length (Vfull) and a truncated form (V307). Mouse challenge studies with Y. pestis strain CO92 showed that vaccination with a combination of RCN-F1 and the truncated V construct (RCN-V307) provided the greatest improvement (P =0.01) in protection against plague over vaccination with RCN-F1 alone. This effect was mediated primarily by anti-F1 and anti-V antibodies and both contributed independently to increased survival of vaccinated mice. [Copyright &y& Elsevier]