An Equine Herpesvirus Type 1 (EHV-1) Ab4 Open Reading Frame 2 Deletion Mutant Provides Immunity and Protection from EHV-1 Infection and Disease

Abstract

Equine herpesvirus type 1 (EHV-1) outbreaks continue to occur despite widely used vaccination. Therefore, development of EHV-1 vaccines providing improved immunity and protection is ongoing. Here, an open reading frame 2 deletion mutant of the neuropathogenic EHV-1 strain Ab4 (Ab4ΔORF2) was tested as a vaccine candidate. Three groups of horses (n = 8 each) were infected intranasally with Ab4ΔORF2 or the parent Ab4 virus or were kept as noninfected controls. Horses infected with Ab4ΔORF2 had reduced fever and nasal virus shedding compared to those infected with Ab4 but mounted similar adaptive immunity dominated by antibody responses. Nine months after the initial infection, all horses were challenged intranasally with Ab4. Previously noninfected horses (control/Ab4) displayed clinical signs, shed large amounts of virus, and developed cell-associated viremia. In contrast, 5/8 or 3/8 horses previously infected with Ab4ΔORF2 or Ab4, respectively, were fully protected from challenge infection as indicated by the absence of fever, clinical disease, nasal virus shedding, and viremia. All of these outcomes were significantly reduced in the remaining, partially protected 3/8 (Ab4ΔORF2/Ab4) and 5/8 (Ab4/Ab4) horses. Protected horses had EHV-1-specific IgG4/7 antibodies prior to challenge infection, and intranasal antibodies increased rapidly postchallenge. Intranasal inflammatory markers were not detectable in protected horses but quickly increased in control/Ab4 horses during the first week after infection. Overall, our data suggest that preexisting nasal IgG4/7 antibodies neutralize EHV-1, prevent viral entry, and thereby protect from disease, viral shedding, and cell-associated viremia. In conclusion, improved protection from challenge infection emphasizes further evaluation of Ab4ΔORF2 as a vaccine candidate. IMPORTANCE Nasal equine herpesvirus type 1 (EHV-1) shedding is essential for virus transmission during outbreaks. Cell-associated viremia is a prerequisite for the most severe disease outcomes, abortion and equine herpesvirus myeloencephalopathy (EHM). Thus, protection from viremia is considered essential for preventing EHM. Ab4ΔORF2 vaccination prevented EHV-1 challenge virus replication in the upper respiratory tract in fully protected horses. Consequently, these neither shed virus nor developed cell-associated viremia. Protection from virus shedding and viremia during challenge infection in combination with reduced virulence at the time of vaccination emphasizes ORF2 deletion as a promising modification for generating an improved EHV-1 vaccine. During this challenge infection, full protection was linked to preexisting local and systemic EHV-1-specific antibodies combined with rapidly increasing intranasal IgG4/7 antibodies and lack of nasal type I interferon and chemokine induction. These host immune parameters may constitute markers of protection against EHV-1 and be utilized as indicators for improved vaccine development and informed vaccination strategies.

Equine Herpesvirus Type 1 Modulates Cytokine and Chemokine Profiles of Mononuclear Cells for Efficient Dissemination to Target Organs

“…We assume that these viral proteins (ORF2 and ORF17) are involved in modulating several pathways in PBMC, including cell signaling, adhesion, and immune pathways, thereby ORF2 and ORF17 gene deletions resulted in reduced virus transfer to EC. Our findings clearly correlate with previous in vivo findings where ORF2, ORF1/ORF71, and ORF1/ORF2 deletion mutants showed respiratory infection, comparable viremia, but no noticeable endothelial cell infection in four different experimental studies in horses [48][49][50]76].…”

“…Experimental infection with ORF1 and ORF71 deletion mutants revealed a brief period of pyrexia, low virus shedding, and decreased cytokine response (IFNα, IL-10, and soluble CD14); however, they had comparable viremia to Ab4-wt [48]. In another study, ORF2 deletion mutant was attenuated; however, it had strong immunogenicity without altering viremia in Icelandic horses [49,76]. Failure to induce T-cell response was suggested as a reason for viremia in both Ab4-wt and mutant viruses infected horses [48][49][50].…”

“…Ab4-wt infection blocks IFNγ release from PBMC and deleting ORF1 gene resulted in release of more cytokines/chemokines which shows the immune-modulating potential of ORF1 in Ab4 strain. ORF2 gene deletion also showed release of more cytokines in both in vitro and in vivo [49,76]. ORF2 gene deletion resulted in expression of higher levels of IL-1β cytokine, which is considered as a host defense against virus infection.…”

Development of a quantitative COVID-19 multiplex assay and its use for serological surveillance in a low SARS-CoV-2 incidence community

“…The ability of the COVID-19 Multiplex Assay to measure both IgG and IgA isotypes will enable further study of the immune response to SARS-CoV-2, including the quantification of antibody responses after vaccination. Furthermore, the COVID-19 Multiplex Assays for IgG and IgA will be valuable tools for measuring mucosal antibody responses due to the improved analytical sensitivity of the bead-based platform compared to conventional ELISA [75] and as shown previously for other respiratory viral pathogens [65,76,77].…”

Application of equine herpesvirus-1 vaccine inactivated by both formaldehyde and binary ethylenimine in equine

“…Efficient vaccination against EHV-1 should induce both humoral and cell-mediated immune responses [ 10 ]. Systemic immunity influences the efficacy of vaccination and its protective effect against EHV-1 [ 11 , 12 ]. The serological condition of mares should involve a low ELISA titer before vaccination, with an inactivated vaccine to give a good immune response [ 13 ], as a poor immune response to vaccination can enhance the severity of disease after infection of vaccinated animals [ 14 ].…”