Prior Heterologous Flavivirus Exposure Results in Reduced Pathogenesis in a Mouse Model of Zika Virus Infection.

The 2015/2016 Zika virus epidemic in South and Central America left the scientific community urgently trying to understand the factors that contribute to Zika virus pathogenesis. Because multiple other flaviviruses are endemic in areas where Zika virus emerged, it is hypothesized that a key to understanding Zika virus disease severity is to study Zika virus infection in the context of prior flavivirus exposure. Human and animal studies have highlighted the idea that having been previously exposed to a different flavivirus may modulate the immune response to Zika virus. However, it is still unclear how prior flavivirus exposure impacts Zika viral burden and disease. In this murine study, we longitudinally examine multiple factors involved in Zika disease, linking viral burden with increased neurological disease severity, weight loss, and inflammation. We show that prior heterologous flavivirus exposure with dengue virus type 2 or 3 or the vaccine strain of yellow fever provides protection from mortality in a lethal Zika virus challenge. However, reduction in viral burden and Zika disease varies depending on the infecting primary flavivirus; with primary Zika virus infection being most protective from Zika virus challenge, followed by dengue virus 2, with yellow fever and dengue virus 3 protecting against mortality but showing more severe disease. This study demonstrates the variation in protective effects of prior flavivirus exposure on Zika virus pathogenesis and identifies distinct relationships between primary flavivirus infection and the potential for Zika virus disease. IMPORTANCE The emergence and reemergence of various vector-borne diseases in recent years highlights the need to understand the mechanisms of protection for each pathogen. In this study, we investigated the impact of prior exposure to Zika virus, dengue virus serotypes 2 or 3, or the vaccine strain of yellow fever on pathogenesis and disease outcomes in a mouse model of Zika virus infection. We found that prior exposure to a heterologous flavivirus was protective from mortality, and to varying degrees, prior flavivirus exposure was protective against neurological disease, weight loss, and severe viral burden during a lethal Zika challenge. Using a longitudinal and cross-sectional study design, we were able to link multiple disease parameters, including viral burden, with neurological disease severity, weight loss, and inflammatory response in the context of flavivirus infection. This study demonstrates a measurable but varied impact of prior flavivirus exposure in modulating flavivirus pathophysiology. Given the cyclic nature of most flavivirus outbreaks, this work will contribute to the forecasting of disease severity for future outbreaks.

mRNA induced expression of human angiotensin-converting enzyme 2 in mice for the study of the adaptive immune response to severe acute respiratory syndrome coronavirus 2.

The novel human coronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a pandemic. Critical to the rapid evaluation of vaccines and antivirals against SARS-CoV-2 is the development of tractable animal models to understand the adaptive immune response to the virus. To this end, the use of common laboratory strains of mice is hindered by significant divergence of the angiotensin-converting enzyme 2 (ACE2), which is the receptor required for entry of SARS-CoV-2. In the current study, we designed and utilized an mRNA-based transfection system to induce expression of the hACE2 receptor in order to confer entry of SARS-CoV-2 in otherwise non-permissive cells. By employing this expression system in an in vivo setting, we were able to interrogate the adaptive immune response to SARS-CoV-2 in type 1 interferon receptor deficient mice. In doing so, we showed that the T cell response to SARS-CoV-2 is enhanced when hACE2 is expressed during infection. Moreover, we demonstrated that these responses are preserved in memory and are boosted upon secondary infection. Importantly, using this system, we functionally identified the CD4+ and CD8+ structural peptide epitopes targeted during SARS-CoV-2 infection in H2b restricted mice and confirmed their existence in an established model of SARS-CoV-2 pathogenesis. We demonstrated that, identical to what has been seen in humans, the antigen-specific CD8+ T cells in mice primarily target peptides of the spike and membrane proteins, while the antigen-specific CD4+ T cells target peptides of the nucleocapsid, membrane, and spike proteins. As the focus of the immune response in mice is highly similar to that of the humans, the identification of functional murine SARS-CoV-2-specific T cell epitopes provided in this study will be critical for evaluation of vaccine efficacy in murine models of SARS-CoV-2 infection.

The role of vaccination route with an adenovirus-vectored vaccine in protection, viral control, and transmission in the SARS-CoV-2/K18-hACE2 mouse infection model.

Introduction: Vaccination is the most effective mechanism to prevent severe COVID-19. However, breakthrough infections and subsequent transmission of SARS-CoV-2 remain a significant problem. Intranasal vaccination has the potential to be more effective in preventing disease and limiting transmission between individuals as it induces potent responses at mucosal sites. Methods: Utilizing a replication-deficient adenovirus serotype 5-vectored vaccine expressing the SARS-CoV-2 RBD (AdCOVID) in homozygous and heterozygous transgenic K18-hACE2, we investigated the impact of the route of administration on vaccine immunogenicity, SARS-CoV-2 transmission, and survival. Results: Mice vaccinated with AdCOVID via the intramuscular or intranasal route and subsequently challenged with SARS-CoV-2 showed that animals vaccinated intranasally had improved cellular and mucosal antibody responses. Additionally, intranasally vaccinated animals had significantly better viremic control, and protection from lethal infection compared to intramuscularly vaccinated animals. Notably, in a novel transmission model, intranasal vaccination reduced viral transmission to naïve co-housed mice compared to intramuscular vaccination. Discussion: Our data provide convincing evidence for the use of intranasal vaccination in protecting against SARS-CoV-2 infection and transmission.

Obesity Enhances Disease Severity in Female Mice Following West Nile Virus Infection.

A rise in adiposity in the United States has resulted in more than 70% of adults being overweight or obese, and global obesity rates have tripled since 1975. Following the 2009 H1N1 pandemic, obesity was characterized as a risk factor that could predict severe infection outcomes to viral infection. Amidst the SARS-CoV-2 pandemic, obesity has remained a significant risk factor for severe viral disease as obese patients have a higher likelihood for developing severe symptoms and requiring hospitalization. However, the mechanism by which obesity enhances viral disease is unknown. In this study, we utilized a diet-induced obesity mouse model of West Nile virus (WNV) infection, a flavivirus that cycles between birds and mosquitoes and incidentally infects both humans and mice. Likelihood for severe WNV disease is associated with risk factors such as diabetes that are comorbidities also linked to obesity. Utilizing this model, we showed that obesity-associated chronic inflammation increased viral disease severity as obese female mice displayed higher mortality rates and elevated viral titers in the central nervous system. In addition, our studies highlighted that obesity also dysregulates host acute adaptive immune responses, as obese female mice displayed significant dysfunction in neutralizing antibody function. These studies highlight that obesity-induced immunological dysfunction begins at early time points post infection and is sustained through memory phase, thus illuminating a potential for obesity to alter the differentiation landscape of adaptive immune cells.

Corticosteroid treatment in COVID-19 modulates host inflammatory responses and transcriptional signatures of immune dysregulation.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of coronavirus disease-2019 (COVID-19), a respiratory disease that varies in severity from mild to severe/fatal. Several risk factors for severe disease have been identified, notably age, male sex, and pre-existing conditions such as diabetes, obesity, and hypertension. Several advancements in clinical care have been achieved over the past year, including the use of corticosteroids (e.g., corticosteroids) and other immune-modulatory treatments that have now become standard of care for patients with acute severe COVID-19. While the understanding of the mechanisms that underlie increased disease severity with age has improved over the past few months, it remains incomplete. Furthermore, the molecular impact of corticosteroid treatment on host response to acute SARS-CoV-2 infection has not been investigated. In this study, a cross-sectional and longitudinal analysis of Ab, soluble immune mediators, and transcriptional responses in young (65 ≤ years) and aged (≥ 65 years) diabetic males with obesity hospitalized with acute severe COVID-19 was conducted. Additionally, the transcriptional profiles in samples obtained before and after corticosteroids became standard of care were compared. The analysis indicates that severe COVID-19 is characterized by robust Ab responses, heightened systemic inflammation, increased expression of genes related to inflammatory and pro-apoptotic processes, and reduced expression of those important for adaptive immunity regardless of age. In contrast, COVID-19 patients receiving steroids did not show high levels of systemic immune mediators and lacked transcriptional indicators of heightened inflammatory and apoptotic responses. Overall, these data suggest that inflammation and cell death are key drivers of severe COVID-19 pathogenesis in the absence of corticosteroid therapy.

The Ability of Zika virus Intravenous Immunoglobulin to Protect From or Enhance Zika Virus Disease.

The closely related flaviviruses, dengue and Zika, cause significant human disease throughout the world. While cross-reactive antibodies have been demonstrated to have the capacity to potentiate disease or mediate protection during flavivirus infection, the mechanisms responsible for this dichotomy are still poorly understood. To understand how the human polyclonal antibody response can protect against, and potentiate the disease in the context of dengue and Zika virus infection we used intravenous hyperimmunoglobulin (IVIG) preparations in a mouse model of the disease. Three IVIGs (ZIKV-IG, Control-Ig and Gamunex®) were evaluated for their ability to neutralize and/or enhance Zika, dengue 2 and 3 viruses in vitro. The balance between virus neutralization and enhancement provided by the in vitro neutralization data was used to predict the IVIG concentrations which could protect or enhance Zika, and dengue 2 disease in vivo. Using this approach, we were able to define the unique in vivo dynamics of complex polyclonal antibodies, allowing for both enhancement and protection from flavivirus infection. Our results provide a novel understanding of how polyclonal antibodies interact with viruses with implications for the use of polyclonal antibody therapeutics and the development and evaluation of the next generation flavivirus vaccines.

Current Flavivirus Research Important for Vaccine Development.

The Flaviviridae family of RNA viruses includes numerous human disease-causing pathogens that largely are increasing in prevalence due to continual climate change, rising population sizes and improved ease of global travel [...].

mRNA induced expression of human angiotensin-converting enzyme 2 in mice for the study of the adaptive immune response to severe acute respiratory syndrome coronavirus 2.

The novel human coronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a pandemic resulting in nearly 20 million infections across the globe, as of August 2020. Critical to the rapid evaluation of vaccines and antivirals is the development of tractable animal models of infection. The use of common laboratory strains of mice to this end is hindered by significant divergence of the angiotensin-converting enzyme 2 (ACE2), which is the receptor required for entry of SARS-CoV-2. In the current study, we designed and utilized an mRNA-based transfection system to induce expression of the hACE2 receptor in order to confer entry of SARS-CoV-2 in otherwise non-permissive cells. By employing this expression system in an in vivo setting, we were able to interrogate the adaptive immune response to SARS-CoV-2 in type 1 interferon receptor deficient mice. In doing so, we showed that the T cell response to SARS-CoV-2 is enhanced when hACE2 is expressed during infection. Moreover, we demonstrated that these responses are preserved in memory and are boosted upon secondary infection. Interestingly, we did not observe an enhancement of SARS-CoV-2 specific antibody responses with hACE2 induction. Importantly, using this system, we functionally identified the CD4+ and CD8+ peptide epitopes targeted during SARS-CoV-2 infection in H2b restricted mice. Antigen-specific CD8+ T cells in mice of this MHC haplotype primarily target peptides of the spike and membrane proteins, while the antigen-specific CD4+ T cells target peptides of the nucleocapsid, membrane, and spike proteins. The functional identification of these T cell epitopes will be critical for evaluation of vaccine efficacy in murine models of SARS-CoV-2. The use of this tractable expression system has the potential to be used in other instances of emerging infections in which the rapid development of an animal model is hindered by a lack of host susceptibility factors.

Immunogenicity and Efficacy of a Recombinant Human Adenovirus Type 5 Vaccine against Zika Virus.

Zika virus (ZIKV) is a significant public health concern due to the pathogen's ability to be transmitted by either mosquito bite or sexual transmission, allowing spread to occur throughout the world. The potential consequences of ZIKV infection to human health, specifically neonates, necessitates the development of a safe and effective Zika virus vaccine. Here, we developed an intranasal Zika vaccine based upon the replication-deficient human adenovirus serotype 5 (hAd5) expressing ZIKV pre-membrane and envelope protein (hAd5-ZKV). The hAd5-ZKV vaccine is able to induce both cell-mediated and humoral immune responses to ZIKV epitopes. Importantly, this vaccine generated CD8+ T cells specific for a dominant ZIKV T cell epitope and is shown to be protective against a ZIKV challenge by using a pre-clinical model of ZIKV disease. We also demonstrate that the vaccine expresses pre-membrane and envelope protein in a confirmation recognized by ZIKV experienced individuals. Our studies demonstrate that this adenovirus-based vaccine expressing ZIKV proteins is immunogenic and protective in mice, and it encodes ZIKV proteins in a conformation recognized by the human antibody repertoire.