Viktoriya Borisevich

Superior efficacy of a recombinant flagellin:H5N1 HA globular head vaccine is determined by the placement of the globular head within flagellin

Transmission of highly pathogenic avian influenza (HPAI) between birds and humans is an ongoing threat that holds potential for the emergence of a pandemic influenza strain. A major barrier to an effective vaccine against avian influenza has been the generally poor immunopotency of many of the HPAI strains coupled with the manufacturing constraints employing conventional methodologies. Fusion of flagellin, a toll-like receptor-5 ligand, to vaccine antigens has been shown to enhance the immune response to the fused antigen in preclinical studies. Here, we have evaluated the immunogenicity and efficacy of a panel of flagellin-based hemagglutinin (HA) globular head fusion vaccines in inbred mice. The HA globular head of these vaccines is derived from the A/Vietnam/1203/04 (VN04; H5N1) HA molecule. We find that replacement of domain D3 of flagellin with the VN04 HA globular head creates a highly effective vaccine that elicits protective HAI titers which protect mice against disease and death in a lethal challenge model.

Inhibition of alphavirus infection in cell culture and in mice with antisense morpholino oligomers

Abstract The genus Alphavirus contains members that threaten human health, both as natural pathogens and as potential biological weapons. Peptide-conjugated phosphorodiamidate morpholino oligomers (PPMO) enter cells readily and can inhibit viral replication through sequence-specific steric blockade of viral RNA. Sindbis virus (SINV) has low pathogenicity in humans and is regularly utilized as a model alphavirus. PPMO targeting the 5′-terminal and AUG translation start site regions of the SINV genome blocked the production of infectious SINV in tissue culture. PPMO designed against corresponding regions in Venezuelan equine encephalitis virus (VEEV) were likewise found to be effective in vitro against several strains of VEEV. Mice treated with PPMO before and after VEEV infection were completely protected from lethal outcome while mice receiving only post-infection PPMO treatment were partially protected. Levels of virus in tissue samples correlated with animal survival. Uninfected mice suffered no apparent ill-effects from PPMO treatment. Thus, PPMO appear promising as candidates for therapeutic development against alphaviruses.

Henipavirus Pathogenesis in Human Respiratory Epithelial Cells

ABSTRACT Hendra virus (HeV) and Nipah virus (NiV) are deadly zoonotic viruses for which no vaccines or therapeutics are licensed for human use. Henipavirus infection causes severe respiratory illness and encephalitis. Although the exact route of transmission in human is unknown, epidemiological studies and in vivo studies suggest that the respiratory tract is important for virus replication. However, the target cells in the respiratory tract are unknown, as are the mechanisms by which henipaviruses can cause disease. In this study, we characterized henipavirus pathogenesis using primary cells derived from the human respiratory tract. The growth kinetics of NiV-Malaysia, NiV-Bangladesh, and HeV were determined in bronchial/tracheal epithelial cells (NHBE) and small airway epithelial cells (SAEC). In addition, host responses to infection were assessed by gene expression analysis and immunoassays. Viruses replicated efficiently in both cell types and induced large syncytia. The host response to henipavirus infection in NHBE and SAEC highlighted a difference in the inflammatory response between HeV and NiV strains as well as intrinsic differences in the ability to mount an inflammatory response between NHBE and SAEC. These responses were highest during HeV infection in SAEC, as characterized by the levels of key cytokines (interleukin 6 [IL-6], IL-8, IL-1α, monocyte chemoattractant protein 1 [MCP-1], and colony-stimulating factors) responsible for immune cell recruitment. Finally, we identified virus strain-dependent variability in type I interferon antagonism in NHBE and SAEC: NiV-Malaysia counteracted this pathway more efficiently than NiV-Bangladesh and HeV. These results provide crucial new information in the understanding of henipavirus pathogenesis in the human respiratory tract at an early stage of infection.

CD4+ T cells provide protection against acute lethal encephalitis caused by Venezuelan equine encephalitis virus

Studying the mechanisms of host survival resulting from viral encephalitis is critical to the development of vaccines. Here we have shown in several independent studies that high dose treatment with neutralizing antibody prior to intranasal infection with Venezuelan equine encephalitis virus had an antiviral effect in the visceral organs and prolonged survival time of infected mice, even in the absence of alphabeta T cells. Nevertheless, antibody treatment did not prevent the development of lethal encephalitis. On the contrary, the adoptive transfer of primed CD4(+) T cells was necessary to prevent lethal encephalitis in mice lacking alphabeta T cell receptor.

Pathogenesis of Hendra and Nipah virus infection in humans

Hendra virus (HeV) and Nipah virus (NiV) are emerging zoonotic viruses that cause severe and often lethal respiratory illness and encephalitis in humans. Henipaviruses can infect a wide range of species and human-to-human transmission has been observed for NiV. While the exact route of transmission in humans is not known, experimental infection in different animal species suggests that infection can be efficiently initiated after respiratory challenge. The limited data on histopathological changes in fatal human cases of HeV and NiV suggest that endothelial cells are an important target during the terminal stage of infection; however, it is unknown where these viruses initially establish infection and how the virus disseminates from the respiratory tract to the central nervous system and other organs. Here we review the current concepts in henipavirus pathogenesis in humans.

The Domestic Ferret (Mustela putorius furo) as a Lethal Infection Model for 3 Species of Ebolavirus

Small-animal models have been developed for several Filoviridae species; however, serial adaptation was required to produce lethal infection. These adapted viruses have sequence changes in several genes, including those that modulate the host immune response. Nonhuman primate models do not require adaptation of filoviruses. Here, we describe lethal models of disease for Bundibugyo, Sudan, and Zaire species of Ebolavirus in the domestic ferret, using wild-type nonadapted viruses. Pathologic features were consistent with disease in primates. Of particular importance, this is the only known small-animal model developed for Bundibugyo and the only uniformly lethal animal model for Bundibugyo.

Prevention of influenza virus shedding and protection from lethal H1N1 challenge using a consensus 2009 H1N1 HA and NA adenovirus vector vaccine.

Vaccines against emerging pathogens such as the 2009 H1N1 pandemic virus can benefit from current technologies such as rapid genomic sequencing to construct the most biologically relevant vaccine. A novel platform (Ad5 [E1-, E2b-]) has been utilized to induce immune responses to various antigenic targets. We employed this vector platform to express hemagglutinin (HA) and neuraminidase (NA) genes from 2009 H1N1 pandemic viruses. Inserts were consensuses sequences designed from viral isolate sequences and the vaccine was rapidly constructed and produced. Vaccination induced H1N1 immune responses in mice, which afforded protection from lethal virus challenge. In ferrets, vaccination protected from disease development and significantly reduced viral titers in nasal washes. H1N1 cell mediated immunity as well as antibody induction correlated with the prevention of disease symptoms and reduction of virus replication. The Ad5 [E1-, E2b-] should be evaluated for the rapid development of effective vaccines against infectious diseases.

Mice Lacking Functional STAT1 Are Highly Susceptible to Lethal Infection with Lassa Virus

ABSTRACT Lassa fever (LF) is a potentially lethal human disease that is caused by the arenavirus Lassa virus (LASV). Annually, around 300,000 infections with up to 10,000 deaths occur in regions of Lassa fever endemicity in West Africa. Here we demonstrate that mice lacking a functional STAT1 pathway are highly susceptible to infection with LASV and develop lethal disease with pathology similar to that reported in humans.

A human lung xenograft mouse model of Nipah virus infection.

Nipah virus (NiV) is a member of the genus Henipavirus (family Paramyxoviridae) that causes severe and often lethal respiratory illness and encephalitis in humans with high mortality rates (up to 92%). NiV can cause Acute Lung Injury (ALI) in humans, and human-to-human transmission has been observed in recent outbreaks of NiV. While the exact route of transmission to humans is not known, we have previously shown that NiV can efficiently infect human respiratory epithelial cells. The molecular mechanisms of NiV-associated ALI in the human respiratory tract are unknown. Thus, there is an urgent need for models of henipavirus infection of the human respiratory tract to study the pathogenesis and understand the host responses. Here, we describe a novel human lung xenograft model in mice to study the pathogenesis of NiV. Following transplantation, human fetal lung xenografts rapidly graft and develop mature structures of adult lungs including cartilage, vascular vessels, ciliated pseudostratified columnar epithelium, and primitive “air” spaces filled with mucus and lined by cuboidal to flat epithelium. Following infection, NiV grows to high titers (107 TCID50/gram lung tissue) as early as 3 days post infection (pi). NiV targets both the endothelium as well as respiratory epithelium in the human lung tissues, and results in syncytia formation. NiV infection in the human lung results in the production of several cytokines and chemokines including IL-6, IP-10, eotaxin, G-CSF and GM-CSF on days 5 and 7 pi. In conclusion, this study demonstrates that NiV can replicate to high titers in a novel in vivo model of the human respiratory tract, resulting in a robust inflammatory response, which is known to be associated with ALI. This model will facilitate progress in the fundamental understanding of henipavirus pathogenesis and virus-host interactions; it will also provide biologically relevant models for other respiratory viruses.

The Synthetic Antiviral Drug Arbidol Inhibits Globally Prevalent Pathogenic Viruses

ABSTRACT Arbidol (ARB) is a synthetic antiviral originally developed to combat influenza viruses. ARB is currently used clinically in several countries but not in North America. We have previously shown that ARB inhibits in vitro hepatitis C virus (HCV) by blocking HCV entry and replication. In this report, we expand the list of viruses that are inhibited by ARB and demonstrate that ARB suppresses in vitro infection of mammalian cells with Ebola virus (EBOV), Tacaribe arenavirus, and human herpesvirus 8 (HHV-8). We also confirm suppression of hepatitis B virus and poliovirus by ARB. ARB inhibited EBOV Zaire Kikwit infection when added before or at the same time as virus infection and was less effective when added 24 h after EBOV infection. Experiments with recombinant vesicular stomatitis virus (VSV) expressing the EBOV Zaire glycoprotein showed that infection was inhibited by ARB at early stages, most likely at the level of viral entry into host cells. ARB inhibited HHV-8 replication to a similar degree as cidofovir. Our data broaden the spectrum of antiviral efficacy of ARB to include globally prevalent viruses that cause significant morbidity and mortality. IMPORTANCE There are many globally prevalent viruses for which there are no licensed vaccines or antiviral medicines. Some of these viruses, such as Ebola virus or members of the arenavirus family, rapidly cause severe hemorrhagic diseases that can be fatal. Other viruses, such as hepatitis B virus or human herpesvirus 8 (HHV-8), establish persistent infections that cause chronic illnesses, including cancer. Thus, finding an affordable, effective, and safe drug that blocks many viruses remains an unmet medical need. The antiviral drug arbidol (ARB), already in clinical use in several countries as an anti-influenza treatment, has been previously shown to suppress the growth of many viruses. In this report, we expand the list of viruses that are blocked by ARB in a laboratory setting to include Ebola virus, Tacaribe arenavirus, and HHV-8, and we propose ARB as a broad-spectrum antiviral drug that may be useful against hemorrhagic viruses.