Utilization of an Eilat Virus-Based Chimera for Serological Detection of Chikungunya Infection.

In December of 2013, chikungunya virus (CHIKV), an alphavirus in the family Togaviridae, was introduced to the island of Saint Martin in the Caribbean, resulting in the first autochthonous cases reported in the Americas. As of January 2015, local and imported CHIKV has been reported in 50 American countries with over 1.1 million suspected cases. CHIKV causes a severe arthralgic disease for which there are no approved vaccines or therapeutics. Furthermore, the lack of a commercially available, sensitive, and affordable diagnostic assay limits surveillance and control efforts. To address this issue, we utilized an insect-specific alphavirus, Eilat virus (EILV), to develop a diagnostic antigen that does not require biosafety containment facilities to produce. We demonstrated that EILV/CHIKV replicates to high titers in insect cells and can be applied directly in enzyme-linked immunosorbent assays without inactivation, resulting in highly sensitive detection of recent and past CHIKV infection, and outperforming traditional antigen preparations

Heterologous Immunization With Defined RNA and Subunit Vaccines Enhances T Cell Responses That Protect Against Leishmania donovani

The rapid generation of strong T cell responses is highly desirable and viral vectors can have potent CD8+ T cell-inducing activity. Immunity to leishmaniasis requires selective T cell responses, with immunization schemes that raise either CD4 or CD8 T cell responses being protective in small animal models. We have defined the leishmaniasis vaccine candidate recombinant fusion antigens, LEISH-F2 and LEISH-F3+, that when formulated in a stable emulsion with a Toll-like receptor (TLR) 4 agonist, induce protective CD4+ T cell responses in animal models as well as providing therapeutic efficacy in canine leishmaniasis and in clinical trials in leishmaniasis patients. We used the genetic sequences of these validated vaccine antigens to design RNA vaccine constructs. Immunization of mice with the RNA replicons induced potent, local innate responses that were surprisingly independent of TLR7 and activated antigen-presenting cells (APC) to prime for extremely potent antigen-specific T helper 1 type responses upon heterologous boosting with either of the subunit vaccines (recombinant antigen with second generation glucopyranosyl lipid A in stable oil-in-water emulsion; SLA-SE). Inclusion of RNA in the immunization schedule also generated MHCI-restricted T cell responses. Immunization with LEISH-F2-expressing RNA vaccine followed later by subunit vaccine afforded protection against challenge with Leishmania donovani. Together, these data indicate the utility of heterologous prime-boost immunization schemes for the induction of potent antigen-specific CD4 and CD8 T cell responses for protection against intracellular pathogens

Chikungunya Virus as Cause of Febrile Illness Outbreak, Chiapas, Mexico, 2014

Since chikungunya virus (CHIKV) was introduced into the Americas in 2013, its geographic distribution has rapidly expanded. Of 119 serum samples collected in 2014 from febrile patients in southern Mexico, 79% were positive for CHIKV or IgM against CHIKV. Sequencing results confirmed CHIKV strains closely related to Caribbean isolates

The evolving SARS-CoV-2 epidemic in Africa: Insights from rapidly expanding genomic surveillance

INTRODUCTION Investment in Africa over the past year with regard to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sequencing has led to a massive increase in the number of sequences, which, to date, exceeds 100,000 sequences generated to track the pandemic on the continent. These sequences have profoundly affected how public health officials in Africa have navigated the COVID-19 pandemic. RATIONALE We demonstrate how the first 100,000 SARS-CoV-2 sequences from Africa have helped monitor the epidemic on the continent, how genomic surveillance expanded over the course of the pandemic, and how we adapted our sequencing methods to deal with an evolving virus. Finally, we also examine how viral lineages have spread across the continent in a phylogeographic framework to gain insights into the underlying temporal and spatial transmission dynamics for several variants of concern (VOCs). RESULTS Our results indicate that the number of countries in Africa that can sequence the virus within their own borders is growing and that this is coupled with a shorter turnaround time from the time of sampling to sequence submission. Ongoing evolution necessitated the continual updating of primer sets, and, as a result, eight primer sets were designed in tandem with viral evolution and used to ensure effective sequencing of the virus. The pandemic unfolded through multiple waves of infection that were each driven by distinct genetic lineages, with B.1-like ancestral strains associated with the first pandemic wave of infections in 2020. Successive waves on the continent were fueled by different VOCs, with Alpha and Beta cocirculating in distinct spatial patterns during the second wave and Delta and Omicron affecting the whole continent during the third and fourth waves, respectively. Phylogeographic reconstruction points toward distinct differences in viral importation and exportation patterns associated with the Alpha, Beta, Delta, and Omicron variants and subvariants, when considering both Africa versus the rest of the world and viral dissemination within the continent. Our epidemiological and phylogenetic inferences therefore underscore the heterogeneous nature of the pandemic on the continent and highlight key insights and challenges, for instance, recognizing the limitations of low testing proportions. We also highlight the early warning capacity that genomic surveillance in Africa has had for the rest of the world with the detection of new lineages and variants, the most recent being the characterization of various Omicron subvariants. CONCLUSION Sustained investment for diagnostics and genomic surveillance in Africa is needed as the virus continues to evolve. This is important not only to help combat SARS-CoV-2 on the continent but also because it can be used as a platform to help address the many emerging and reemerging infectious disease threats in Africa. In particular, capacity building for local sequencing within countries or within the continent should be prioritized because this is generally associated with shorter turnaround times, providing the most benefit to local public health authorities tasked with pandemic response and mitigation and allowing for the fastest reaction to localized outbreaks. These investments are crucial for pandemic preparedness and response and will serve the health of the continent well into the 21st century

An Intramuscular DNA Vaccine for SARS-CoV-2 Decreases Viral Lung Load but Not Lung Pathology in Syrian Hamsters

The 2019 novel coronavirus, SARS-CoV-2, first reported in December 2019, has infected over 102 million people around the world as of February 2021 and thus calls for rapid development of safe and effective interventions, namely vaccines. In our study, we evaluated a DNA vaccine against SARS-CoV-2 in the Syrian hamster model. Hamsters were vaccinated with a DNA-plasmid encoding the SARS-CoV-2 full length spike open reading frame (ORF) to induce host cells to produce spike protein and protective immune responses before exposure to infectious virus. We tested this vaccine candidate by both intranasal (IN) and intramuscular (IM) routes of administration and complexing with and without an in vivo delivery reagent. Hamsters receiving prime-boost-boost IM-only vaccinations recovered body weight quicker, had decreased lung viral loads, and increased SARS-CoV-2-specific antibody titers compared to control vaccinated animals but, surprisingly, lung pathology was as severe as sham vaccinated controls. The IM/IN combination group showed no efficacy in reducing lung virus titers or pathology. With increasing public health need for rapid and effective interventions, our data demonstrate that in some vaccine contexts, significant antibody responses and decreased viral loads may not be sufficient to prevent lung pathology

An RNA Vaccine Protects against Crimean-Congo Hemorrhagic Fever Virus Infection in Mice

Crimean-Congo Hemorrhagic Fever Virus (CCHFV) is commonly transmitted by ticks and has a wide geographic distribution, being endemic throughout southern and eastern Europe, the Middle East, Asia, and Africa. Due to climate change, this range is even expanding northward. CCHFV causes a hemorrhagic disease characterized by initial non-specific illness (fever, muscle pains, nausea) followed by severe internal and external bleeding with a case fatality rate of 5-60%. As there is no licensed therapeutics or vaccines, there is great need for highly effective countermeasures. Recently, we have developed a self-replicating RNA-based vaccine which expresses two of the viral proteins: the CCHFV nucleoprotein (repNP) and glycoprotein precursor (repGPC) and is highly protective against a lethal viral challenge in mice. We found that vaccination with this vaccine prevented disease and conferred 100% survival against a lethal CCHFV infection in mice. This vaccine significantly stimulated both antibody-producing B-cells and T-cells which kill infected cells. Our repNP vaccination primarily stimulated B-cells to produce anti-NP antibodies while repGPC primarily stimulated the T-cells. To confirm whether B-cells or T-cells are most responsible for protection, we vaccinated mice lacking B- or T-cells. While mice lacking T-cells survived, B-cell deficient mice only had ~40% survival indicating that the B-cells and antibodies are essential to confer protection. Currently, this vaccine is undergoing further preclinical testing in preparation for human clinical trials while ongoing studies are continuing to investigate how these antibodies protect against CCHFV so that we may improve additional and much needed CCHFV therapeutics. This research was funded by the Intramural Research Program, NIAID, NIH

A replicating RNA vaccine confers protection in a rhesus macaque model of Crimean-Congo hemorrhagic fever

Abstract Crimean-Congo hemorrhagic fever (CCHF) is a tick-borne febrile illness with a wide geographic distribution. In recent years the geographic range of Crimean-Congo hemorrhagic fever virus (CCHFV) and its tick vector have increased, placing an increasing number of people at risk of CCHFV infection. Currently, there are no widely available vaccines, and although the World Health Organization recommends ribavirin for treatment, its efficacy is unclear. Here we evaluate a promising replicating RNA vaccine in a rhesus macaque (Macaca mulatta) model of CCHF. This model provides an alternative to the established cynomolgus macaque model and recapitulates mild-to-moderate human disease. Rhesus macaques infected with CCHFV consistently exhibit viremia, detectable viral RNA in a multitude of tissues, and moderate pathology in the liver and spleen. We used this model to evaluate the immunogenicity and protective efficacy of a replicating RNA vaccine. Rhesus macaques vaccinated with RNAs expressing the CCHFV nucleoprotein and glycoprotein precursor developed robust non-neutralizing humoral immunity against the CCHFV nucleoprotein and had significant protection against the CCHFV challenge. Together, our data report a model of CCHF using rhesus macaques and demonstrate that our replicating RNA vaccine is immunogenic and protective in non-human primates after a prime-boost immunization

Replicating RNA vaccination elicits an unexpected immune response that efficiently protects mice against lethal Crimean-Congo hemorrhagic fever virus challenge

Summary: Background: Crimean-Congo hemorrhagic fever virus is the cause of a severe hemorrhagic fever with cases reported throughout a wide-geographic region. Spread by the bite of infected ticks, contact with infected livestock or in the health care setting, disease begins as a non-specific febrile illness that can rapidly progress to hemorrhagic manifestations. Currently, there are no approved vaccines and antivirals such as ribavirin have unclear efficacy. Thus treatment is mostly limited to supportive care. Methods: In this report we evaluated an alphavirus-based replicon RNA vaccine expressing either the CCHFV nucleoprotein or glycoprotein precursor in a stringent, heterologous lethal challenge mouse model. Findings: Vaccination with the RNA expressing the nucleoprotein alone could confer complete protection against clinical disease, but vaccination with a combination of both the nucleoprotein and glycoprotein precursor afforded robust protection against disease and viral replication. Protection from lethal challenge required as little as a single immunization with 100ng of RNA. Unexpectedly, analysis of the immune responses elicited by the vaccine components showed that vaccination resulted in antibodies against the internal viral nucleoprotein and cellular immunity against the virion-exposed glycoproteins. Interpretation: Cumulatively this vaccine conferred robust protection against Crimean-Congo hemorrhagic fever virus and supports continued development of this vaccine candidate. Funding: This research was supported by the Intramural Research Program of the NIAID/NIH and HDT Bio