Impact of Menglà Virus Proteins on Human and Bat Innate Immune Pathways.

Menglà virus (MLAV), identified in Rousettus bats, is a phylogenetically distinct member of the family Filoviridae Because the filoviruses Ebola virus (EBOV) and Marburg virus (MARV) modulate host innate immunity, MLAV VP35, VP40, and VP24 proteins were compared with their EBOV and MARV homologs for innate immune pathway modulation. In human and Rousettus cells, MLAV VP35 behaved like EBOV and MARV VP35s, inhibiting virus-induced activation of the interferon beta (IFN-ß) promoter and interferon regulatory factor 3 (IRF3) phosphorylation. MLAV VP35 also interacted with PACT, a host protein engaged by EBOV VP35 to inhibit RIG-I signaling. MLAV VP35 also inhibits PKR activation. MLAV VP40 was demonstrated to inhibit type I IFN-induced gene expression in human and bat cells. It blocked STAT1 tyrosine phosphorylation induced either by type I IFN or overexpressed Jak1, paralleling MARV VP40. MLAV VP40 also inhibited virus-induced IFN-ß promoter activation, a property shared by MARV VP40 and EBOV VP24. A Jak kinase inhibitor did not recapitulate this inhibition in the absence of viral proteins. Therefore, inhibition of Jak-STAT signaling is insufficient to explain inhibition of IFN-ß promoter activation. MLAV VP24 did not inhibit IFN-induced gene expression or bind karyopherin α proteins, properties of EBOV VP24. MLAV VP24 differed from MARV VP24 in that it failed to interact with Keap1 or activate an antioxidant response element reporter gene due to the absence of a Keap1-binding motif. These functional observations support a closer relationship of MLAV to MARV than to EBOV but also are consistent with MLAV belonging to a distinct genus.IMPORTANCE EBOV and MARV, members of the family Filoviridae, are highly pathogenic zoonotic viruses that cause severe disease in humans. Both viruses use several mechanisms to modulate the host innate immune response, and these likely contribute to the severity of disease. Here, we demonstrate that MLAV, a filovirus newly discovered in a bat, suppresses antiviral type I interferon responses in both human and bat cells. Inhibitory activities are possessed by MLAV VP35 and VP40, which parallels how MARV blocks IFN responses. However, whereas MARV activates cellular antioxidant responses through an interaction between its VP24 protein and host protein Keap1, MLAV VP24 lacks a Keap1-binding motif and fails to activate this cytoprotective response. These data indicate that MLAV possesses immune-suppressing functions that could facilitate human infection. They also support the placement of MLAV in a different genus than either EBOV or MARV.

Therapeutics for filovirus infection: traditional approaches and progress towards in silico drug design.

INTRODUCTION: Ebolaviruses and marburgviruses cause severe and often lethal human hemorrhagic fevers. As no FDA-approved therapeutics are available for these infections, efforts to discover new therapeutics are important, especially because these pathogens are considered biothreats and emerging infectious diseases. All methods for discovering new therapeutics should be considered, including compound library screening in vitro against virus and in silico structure-based drug design, where possible, if sufficient biochemical and structural information is available. AREAS COVERED: This review covers the structure and function of filovirus proteins, as they have been reported to date, as well as some of the current antiviral screening approaches. The authors discuss key studies mapping small-molecule modulators that were found through library and in silico screens to potential sites on viral proteins or host proteins involved in virus trafficking and pathogenesis. A description of ebolavirus and marburgvirus diseases and available animal models is also presented. EXPERT OPINION: To discover novel therapeutics with potent efficacy using sophisticated computational methods, more high-resolution crystal structures of filovirus proteins and more details about the protein functions and host interaction will be required. Current compound screening efforts are finding active antiviral compounds, but an emphasis on discovery research to investigate protein structures and functions enabling in silico drug design would provide another avenue for finding antiviral molecules. Additionally, targeting of protein-protein interactions may be a future avenue for drug discovery since disrupting catalytic sites may not be possible for all proteins.

Lethality and pathogenesis of airborne infection with filoviruses in A129 α/ß -/- interferon receptor-deficient mice.

Normal immunocompetent mice are not susceptible to non-adapted filoviruses. There are therefore two strategies available to establish a murine model of filovirus infection: adaptation of the virus to the host or the use of genetically modified mice that are susceptible to the virus. A number of knockout (KO) strains of mice with defects in either their adaptive or innate immunity are susceptible to non-adapted filoviruses. In this study, A129 α/ß -/- interferon receptor-deficient KO mice, strain A129 IFN-α/ß -/-, were used to determine the lethality of a range of filoviruses, including Lake Victoria marburgvirus (MARV), Zaire ebolavirus (ZEBOV), Sudan ebolavirus (SEBOV), Reston ebolavirus (REBOV) and Côte d'Ivoire ebolavirus (CIEBOV), administered by using intraperitoneal (IP) or aerosol routes of infection. One hundred percent mortality was observed in all groups of KO mice that were administered with a range of challenge doses of MARV and ZEBOV by either IP or aerosol routes. Mean time to death for both routes was dose-dependent and ranged from 5.4 to 7.4 days in the IP injection challenge, and from 10.2 to 13 days in the aerosol challenge. The lethal dose (50 % tissue culture infective dose, TCID(50)) of ZEBOV for KO mice was <1 TCID(50) ml(-1) when administered by either the IP or aerosol route of infection; for MARV the lethal dose was <1 TCID(50) ml(-1) by the IP route of infection and <10 TCID(50) ml(-1) by the aerosol route. In contrast, there was no mortality after infection with SEBOV or REBOV by either IP or aerosol routes of infection; all the mice lost weight (~15 % loss of group mean body weight with SEBOV and ~7 % with REBOV) but recovered to their original weights by day 14 post-challenge. There was no mortality in mice administered with CIEBOV via the IP route of infection and no clinical signs of infection were observed. The progression of disease was faster following infection with ZEBOV than with MARV but ultimately both viruses caused widespread infection with high titres of the infectious viruses in multiple organs. Histopathological observations were consistent with other animal models and showed widespread organ damage. This study suggests that MARV and ZEBOV are more virulent when administered via the IP route rather than by aerosol infection, although both are highly virulent by either route. The KO mouse may provide a useful model to test potential antiviral therapeutics against wild-type filoviruses.

Filoviruses and the balance of innate, adaptive, and inflammatory responses.

The Filoviruses Marburg virus and Ebola virus are among the deadliest of human pathogens, causing fulminant hemorrhagic fevers typified by overmatched specific immune responses and profuse inflammatory responses. Keys to both vaccination and treatment may reside, first, in the understanding of immune dysfunctions that parallel Filoviral disease and, second, in devising ways to redirect and restore normal immune function as well as to mitigate inflammation. Here, we describe how Filoviral infections may subvert innate immune responses through perturbances of dendritic cells and neutrophils, with particular emphasis on the downstream effects on adaptive immunity and inflammation. We suggest that pivotal events may be subject to therapeutic intervention as Filoviruses encounter immune processes.

The role of reverse genetics systems in determining filovirus pathogenicity.

The family Filoviridae is comprised of two genera: Marburgvirus and Ebolavirus. To date minigenome systems have been developed for two Ebola viruses (Reston ebolavirus and Zaire ebolavirus [ZEBOV]) as well as for Lake Victoria marburgvirus, the sole member of the Marburgvirus genus. The use of these minigenome systems has helped characterize functions for many viral proteins in both genera and have provided valuable insight towards the development of an infectious clone system in the case of ZEBOV. The recent development of two such infectious clone systems for ZEBOV now allow effective strategies for experimental mutagenesis to study the biology and pathogenesis of one of the most lethal human pathogens.