Vaccinia viruses with mutations in the E3L gene as potential replication-competent, attenuated vaccines: Scarification vaccination

Smallpox was declared eradicated in 1980 after an intensive vaccination program using different strains of vaccinia virus (VACV; Poxviridae). VACV strain IOC (VACV-IOC) was the seed strain of the smallpox vaccine manufactured by the major vaccine producer in Brazil during the smallpox eradication program. However, little is known about the biological and immunological features as well as the phylogenetic relationships of this first-generation vaccine. In this work, we present a comprehensive characterization of two clones of VACV-IOC. Both clones had low virulence in infected mice and induced a protective immune response against a lethal infection comparable to the response of the licensed vaccine ACAM2000 and the parental strain VACV-IOC. Full-genome sequencing revealed the presence of several fragmented virulence genes that probably are nonfunctional, e.g., F1L, B13R, C10L, K3L, and C3L. Most notably, phylogenetic inference supported by the structural analysis of the genome ends provides evidence of a novel, independent cluster in VACV phylogeny formed by VACV-IOC, the Brazilian field strains Cantagalo (CTGV) and Serro 2 viruses, and horsepox virus, a VACV-like virus supposedly related to an ancestor of the VACV lineage. Our data strongly support the hypothesis that CTGV-like viruses represent feral VACV that evolved in parallel with VACV-IOC after splitting from a most recent common ancestor, probably an ancient smallpox vaccine strain related to horsepox virus. Our data, together with an interesting historical investigation, revisit the origins of VACV and propose new evolutionary relationships between ancient and extant VACV strains, mainly horsepox virus, VACV-IOC/CTGV-like viruses, and Dryvax strain. IMPORTANCEFirst-generation vaccines used to eradicate smallpox had rates of adverse effects that are not acceptable by current health care standards. Moreover, these vaccines are genetically heterogeneous and consist of a pool of quasispecies of VACV. Therefore, the search for new-generation smallpox vaccines that combine low pathogenicity, immune protection, and genetic homogeneity is extremely important. In addition, the phylogenetic relationships and origins of VACV strains are quite nebulous. We show the characterization of two clones of VACV-IOC, a unique smallpox vaccine strain that contributed to smallpox eradication in Brazil. The immunogenicity and reduced virulence make the IOC clones good options for alternative second-generation smallpox vaccines. More importantly, this study reveals the phylogenetic relationship between VACV-IOC, feral VACV established in nature, and the ancestor-like horsepox virus. Our data expand the discussion on the origins and evolutionary connections of VACV lineages. S mallpox, caused by variola virus (genus Orthopoxvirus, family Poxviridae), accounted for millions of deaths throughout human history. The disease was declared eradicated in 1980 after a worldwide program implemented by the WHO involving strict surveillance and intensive vaccination using vaccinia vi...

Section: Discussionmentioning

confidence: 83%

Genomic Analysis, Phenotype, and Virulence of the Historical Brazilian Smallpox Vaccine Strain IOC: Implications for the Origins and Evolutionary Relationships of Vaccinia Virus

Medaglia

Moussatché

Nitsche

et al. 2015

“…Unchecked accumulation of dsRNA may also enhance the immune response through the activation of PKR and other dsRNA-sensing pathways. This approach has had success in the vaccinia virus (VACV) system, where a VACV lacking E3L (48,49) provides a useful comparison. Similar to the PKRinhibitory genes of cytomegaloviruses (26)(27)(28)(29)(30)(31)(32)(33)50), vaccinia virus E3L binds to and sequesters dsRNA and thereby prevents activation of type I interferon-induced PKR and oligoadenylate synthetase (51).…”

Section: Discussionmentioning

confidence: 99%

Vaccination with a Live Attenuated Cytomegalovirus Devoid of a Protein Kinase R Inhibitory Gene Results in Reduced Maternal Viremia and Improved Pregnancy Outcome in a Guinea Pig Congenital Infection Model

Schleiss

Bierle

Swanson

et al. 2015

Development of a vaccine to prevent congenital cytomegalovirus infection is a major public health priority. Live vaccines attenuated through mutations targeting viral mechanisms responsible for evasion of host defense may be both safe and efficacious. Safety and vaccine efficacy were evaluated using a guinea pig cytomegalovirus (GPCMV) model. Recombinant GPCMV with a targeted deletion of gp145 (designated ⌬145), a viral protein kinase R (PKR) inhibitor, was generated. Attenuation was evaluated following inoculation of 10 7 PFU of ⌬145 or parental virus into guinea pigs immunosuppressed with cyclophosphamide. Efficacy was evaluated by immunizing GPCMV-naive guinea pigs twice with either 10 5 or 10 6 PFU of ⌬145, establishing pregnancy, and challenging the guinea pigs with salivary gland-adapted GPCMV. The immune response, maternal viral load, pup mortality, and congenital infection rates in the vaccine and control groups were compared. ⌬145 was substantially attenuated for replication in immunocompromised guinea pigs. Vaccination with ⌬145 induced enzyme-linked immunosorbent assay (ELISA) and neutralizing antibody levels comparable to those achieved in natural infection. In the higher-and lower-dose vaccine groups, pup mortality was reduced to 1/24 (4%) and 4/29 (14%) pups, respectively, whereas it was 26/31 (81%) in unvaccinated control pups (P < 0.0001 for both groups versus the control group). Congenital infection occurred in 20/31 (65%) control pups but only 8/24 (33%) pups in the group vaccinated with 10 6 PFU (P < 0.05). Significant reductions in the magnitude of maternal DNAemia and pup viral load were noted in the vaccine groups compared to those in the controls. Deletion of a GPCMV genome-encoded PKR inhibitor results in a highly attenuated virus that is immunogenic and protective as a vaccine against transplacental infection. Infection with human cytomegalovirus (HCMV) causes considerable morbidity and occasional mortality in immunocompromised solid organ transplant and hematopoietic stem cell transplant patients, HIV-infected individuals, and newborns that acquire infection in utero (1, 2). Due to the lifelong morbidity associated with congenital CMV infection, a preconception vaccine capable of preventing virus transmission to the fetus would provide a highly cost-effective public health advance (3). Unfortunately, the lack of clear immunological correlates of protective immunity has hampered development of an HCMV vaccine. In spite of this uncertainty, there is evidence that virus-neutralizing antibody responses targeting viral envelope glycoproteins, as well as cellular immune responses (CD4 ϩ and CD8 ϩ ) targeting multiple structural and regulatory proteins, play important roles in protection against acquisition and reactivation of infection (4-7).

“…Besides the threat posed to human and animal health by poxviruses, there is also a considerable effort under way to develop recombinant poxvirus-based vaccine vectors. Among such vectors, vaccinia virus E3L mutants are being investigated as a vaccine platform due to their greater safety profile in comparison to wild-type vaccinia virus (18,41). Moreover, the use of myxoma virus as an oncolytic agent for the treatment of cancer is being studied (23).…”

mentioning

confidence: 99%

Comparative Analysis of Poxvirus Orthologues of the Vaccinia Virus E3 Protein: Modulation of Protein Kinase R Activity, Cytokine Responses, and Virus Pathogenicity

Myskiw

Arsenio

Hammett

et al. 2011

Poxviruses are important human and animal pathogens that have evolved elaborate strategies for antagonizing host innate and adaptive immunity. The E3 protein of vaccinia virus, the prototypic member of the orthopoxviruses, functions as an inhibitor of innate immune signaling and is essential for vaccinia virus replication in vivo and in many human cell culture systems. However, the function of orthologues of E3 expressed by poxviruses of other genera with different host specificity remains largely unknown. In the present study, we characterized the E3 orthologues from sheeppox virus, yaba monkey tumor virus, swinepox virus, and myxoma virus for their ability to modulate protein kinase R (PKR) function, cytokine responses and virus pathogenicity. We found that the E3 orthologues of myxoma virus and swinepox virus could suppress PKR activation and interferon (IFN)-induced antiviral activities and restore the host range function of E3 in HeLa cells. In contrast, the E3 orthologues from sheeppox virus and yaba monkey tumor virus were unable to inhibit PKR activation. While the sheeppox orthologue was unable to restore the host range function of E3, the yaba monkey tumor virus orthologue partially restored E3-deficient vaccinia virus replication in HeLa cells, correlated with its ability to suppress IFN-induced antiviral activities. Moreover, poxvirus E3 orthologues show varying ability to inhibit the induction of antiviral and proinflammatory cytokines. Despite these in vitro results, none of the E3 orthologues tested was capable of restoring pathogenicity to E3-deficient vaccinia virus in vivo.