MECHANISMS OF TYPE-I IFN INHIBITION: EQUINE HERPESVIRUS-1 ESCAPE FROM THE ANTIVIRAL EFFECT OF TYPE-1 INTERFERON RESPONSE IN HOST CELL

Equine herpesvirus-1 (EHV-1) is one of the most important and prevalent viral pathogens of horses causing a major threat to the equine industry throughout most of the world. EHV-1 primarily causes respiratory disease but viral spread to distant organs enables the development of more severe sequelae; abortion and neurologic disease. In order to produce disease, EHV-1 has to overcome the innate barrier of the type-I interferon (IFN) system in host cells. However, the underlying mechanisms employed by EHV-1 to circumvent the type-I IFN response in host cells are not well understood. In this project study, using molecular techniques, we explored how EHV-1 is able to escape the type-I IFN response in host cells during infection. We also investigated whether EHV-4, a closely related but less pathogenic virus, has similar effects on type-I IFN as a clue to understanding how widespread IFN suppressive function is found among equine alphaherpesviruses. Our data showed that inhibition of the type-I IFN response in host cells is not a function of neuropathogenicity of EHV-1 strains. However, a reduced type-I IFN response correlated with pathogenicity as EHV-4, unlike EHV-1, was unable to down-regulate the type-I IFN response in equine endothelial cells (EECs). Investigation of the mechanisms employed by EHV-1 to suppress type-I IFN revealed that the virus sequentially prevented outside-in signaling events that lead to type-I IFN production. Specifically, EHV-1 blocked the expression of Toll-like receptors (TLR) 3 and TLR4 at 6 hours post-infection (hpi) and 12 hpi. EHV-1 also prevented the transcription of IRF7 and IRF9 at different time-points during infection. The virus also perturbed the JAK-STAT signaling pathway by negatively regulating the cellular levels of TYK2 and phosphorylation-mediated activation of STAT2 molecules. Immunofluorescence data revealed that during infection, EHV-1 was able to sequester STAT2 molecules from nuclear translocation. This may be a limiting step preventing the formation of interferon- stimulated gene factor 3 (ISGF3) whose nuclear translocation is required to transactivate interferon-stimulated genes (ISGs) including IRF7. Further investigation showed that unlike EHV-1, EHV-4 only interfered with phosphorylation-mediated activated STAT1 and STAT2 molecules at 3 and 6 hpi. EHV-4 was unable to block TLR3/4 and IRF7/9 mRNA expression at any time-point. Intriguingly, while viral late gene of EHV-1 mediates inhibition of STAT phosphorylation, our data showed that for EHV-4, a virus late gene did not mediate the inhibition of STAT phosphorylation. The findings from this study help illuminate how EHV-1 strategically interferes with limiting steps required for type-I IFN response in host cells to promote pathology. Our data also strengthen the hypothesis that the ability to shut off host factors required for type-I IFN production might be directly related to the degree of pathogenicity of the EHV subtypes

Equine Influenza Virus and Vaccines

Equine influenza virus (EIV) is a constantly evolving viral pathogen that is responsible for yearly outbreaks of respiratory disease in horses termed equine influenza (EI). There is currently no evidence of circulation of the original H7N7 strain of EIV worldwide; however, the EIV H3N8 strain, which was first isolated in the early 1960s, remains a major threat to most of the world’s horse populations. It can also infect dogs. The ability of EIV to constantly accumulate mutations in its antibody-binding sites enables it to evade host protective immunity, making it a successful viral pathogen. Clinical and virological protection against EIV is achieved by stimulation of strong cellular and humoral immunity in vaccinated horses. However, despite EI vaccine updates over the years, EIV remains relevant, because the protective effects of vaccines decay and permit subclinical infections that facilitate transmission into susceptible populations. In this review, we describe how the evolution of EIV drives repeated EI outbreaks even in horse populations with supposedly high vaccination coverage. Next, we discuss the approaches employed to develop efficacious EI vaccines for commercial use and the existing system for recommendations on updating vaccines based on available clinical and virological data to improve protective immunity in vaccinated horse populations. Understanding how EIV biology can be better harnessed to improve EI vaccines is central to controlling EI

A bivalent live-attenuated vaccine for the prevention of equine influenza virus

Vaccination remains the most effective approach for preventing and controlling equine influenza virus (EIV) in horses. However, the ongoing evolution of EIV has increased the genetic and antigenic differences between currently available vaccines and circulating strains, resulting in suboptimal vaccine efficacy. As recommended by the World Organization for Animal Health (OIE), the inclusion of representative strains from clade 1 and clade 2 Florida sublineages of EIV in vaccines may maximize the protection against presently circulating viral strains. In this study, we used reverse genetics technologies to generate a bivalent EIV live-attenuated influenza vaccine (LAIV). We combined our previously described clade 1 EIV LAIV A/equine/Ohio/2003 H3N8 (Ohio/03 LAIV) with a newly generated clade 2 EIV LAIV that contains the six internal genes of Ohio/03 LAIV and the HA and NA of A/equine/Richmond/1/2007 H3N8 (Rich/07 LAIV). The safety profile, immunogenicity, and protection efficacy of this bivalent EIV LAIV was tested in the natural host, horses. Vaccination of horses with the bivalent EIV LAIV, following a prime-boost regimen, was safe and able to confer protection against challenge with clade 1 (A/equine/Kentucky/2014 H3N8) and clade 2 (A/equine/Richmond/2007) wild-type (WT) EIVs, as evidenced by a reduction of clinical signs, fever, and virus excretion. This is the first description of a bivalent LAIV for the prevention of EIV in horses that follows OIE recommendations. In addition, since our bivalent EIV LAIV is based on the use of reverse genetics approaches, our results demonstrate the feasibility of using the backbone of clade 1 Ohio/03 LAIV as a master donor virus (MDV) for the production and rapid update of LAIVs for the control and protection against other EIV strains of epidemiological relevance to horses

The effect of equine herpesvirus type 4 on type-I interferon signaling molecules

Equine herpesvirus type 4 (EHV-4) is mildly pathogenic but is a common cause of respiratory disease in horses worldwide. We previously demonstrated that unlike EHV-1, EHV-4 is not a potent inducer of type-I IFN and does not suppress that IFN response, especially during late infection, when compared to EHV-1 infection in equine endothelial cells (EECs). Here, we investigated the impact of EHV-4 infection in EECs on type-I IFN signaling molecules at 3, 6, and 12 hpi. Findings from our study revealed that EHV-4 did not induce nor suppress TLR3 and TLR4 expression in EECs at all the studied time points. EHV-4 was able to induce variable amounts of IRF7 and IRF9 in EECs with no evidence of suppressive effect on these important transcription factors of IFN-α/β induction. Intriguingly, EHV-4 did interfere with the phosphorylation of STAT1/STAT2 at 3 hpi and 6 hpi, less so at 12 hpi. An active EHV-4 viral gene expression was required for the suppressive effect of EHV-4 on STAT1/STAT2 phosphorylation during early infection. One or more early viral genes of EHV-4 are involved in the suppression of STAT1/STAT2 phosphorylation observed during early time points in EHV-4-infected EECs. The inability of EHV-4 to significantly down-regulate key molecules of type-I IFN signaling may be related to the lower severity of pathogenesis when compared with EHV-1. Harnessing this knowledge may prove useful in controlling future outbreaks of the disease

Serotypes, antimicrobial profiles, and public health significance of Salmonella from camels slaughtered in Maiduguri central abattoir, Nigeria

Aim: This study aimed at determining the serotypes, antimicrobial profiles, and public health importance of Salmonella strains from camels slaughtered at Maiduguri central abattoir, Nigeria. Materials and Methods: Two hundred samples were obtained from camel comprising of intestines, feces, liver, and spleen (n=50 each). Non-lactose fermenting dark center Salmonella colonies were identified using standard biochemical techniques, serotyped and subjected to antimicrobial susceptibility test using minimum inhibition concentration method. Results: Out of the 200 samples collected, 17 were Salmonella positive (spleen=7, intestine=6, feces=3, and liver=1) with a prevalence of 8.5%. Five serotypes comprising Salmonella Eko, 7 (3.5%), Salmonella Uganda, 4 (2.0%), Salmonella Amager, 2 (1.0%), Salmonella Westhampton, 2 (1.0%), and Salmonella Give, 2 (1.0%) were incriminated. Majority of the serotypes were sensitive to the antimicrobials, but one Salmonella Amager exhibited resistance to streptomycin, and one each of Salmonella Uganda and Salmonella Eko were resistant to sulfamethoxazole. Conclusion: This study revealed the prevalence and the antibiotic resistance profile of newly emerging Salmonella from camels in the northeast of Nigeria, which can serve as a means for the transmission of Salmonella to human. Therefore, there is a need for the establishment of national Salmonella surveillance and control programs

Immunogenicity and Protective Efficacy of an Intranasal Live-attenuated Vaccine Against SARS-CoV-2

SUMMARY Global deployment of an effective and safe vaccine is necessary to curtail the coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Here, we evaluated a Newcastle disease virus (NDV)-based vectored-vaccine in mice and hamsters for its immunogenicity, safety and protective efficacy against SARS-CoV-2. Intranasal administration of recombinant (r)NDV-S vaccine expressing spike (S) protein of SARS-CoV-2 to mice induced high levels of SARS-CoV-2-specific neutralizing immunoglobulin A (IgA) and IgG2a antibodies and T cell-mediated immunity. Hamsters immunised with two doses of vaccine showed complete protection from lung infection, inflammation, and pathological lesions following SARS-CoV-2 challenge. Importantly, administration of two doses of intranasal rNDV-S vaccine significantly reduced the SARS-CoV-2 shedding in nasal turbinate and lungs in hamsters. Collectively, intranasal vaccination has the potential to control infection at the site of inoculation, which should prevent both clinical disease and virus transmission to halt the spread of the COVID-19 pandemic