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Filoviridae
The virus family Filoviridae is comprised of the Marburg virus and the four
strains of the Ebola virus: Zaire, Sudan, Reston, and Ivory Coast. Of the four
Ebola viruses, the Reston strain is the only one that does not cause disease
in humans. A definitive reservoir for the viruses has not been fully determined,
though Marburg virus was recently found in fruit bats. Death rates for Marburg
and Ebola Zaire can be as high as 90%. The viruses are spread through
contact with bodily fluids such as blood, vomit, and excrement. Humans can
be infected by both other infected humans and infected primates. Initial
symptoms include fever, chills, vomiting, and muscle aches. Within ten days
most victims develop hypotension, exhibit hemorrhaging, and die from organ
failure and shock.
The pathology of filovirus infection is one of utter destruction of the host’s
body. Once the infectious virions enter through openings in the skin or mucus
membranes, they bind to target cells and enter via endocytosis. Early targets
of the virus include monocytes, macrophages, and dendritic cells, which carry
the infecting filovirus from the site of infection to the lymph nodes. From there,
the virus spreads throughout the lymph system and eventually makes its way
to the vascular system and target organs.
During the beginning stage of infection the virus overwhelms the immune
system. The virus first destroys main components of the innate immune
system via apoptosis, including the monocytes, macrophages, and natural
killer cells. These cells are involved in the initial recognition of cells infected
with foreign antigens and their quick response is needed to halt viral
replication and spread. Filoviruses also suppress the type I Interferon
response. The type I IFNs are molecules that are part of the early signaling
cascade that alerts the body to the infection. The IFNs bind to the IFN
receptor and through a series of activations turns on transcription factors
STAT1 and STAT2 by phosphorylation. These transcription factors then
activate the transcription of genes involved in the IFN response. Studies have
shown Marbug inhibits the phosphorylation of STAT1 and STAT2 leading to
inhibition of the IFN response. Ebola seems to inhibit phosphorylation and
activation of the transcription factor IRF-3 via its protein VP35. IRF-3 is
involved in activating the IFN-β promoter, thus its inhibition by VP35 leads to
inhibition of the IFN-β gene. Taking out the innate immune response allows
the viral infection to spread undetected.
The Ebola VP35 protein has also been shown to inhibit the antiviral response
after interferon induction by preventing activation of the double stranded
RNA-dependent protein kinase (PKR). As stated before, the VP35 protein is
able to inhibit phosphorylation and activation of IRF-3, which in turn inhibits
IFN-β. VP35 can also inhibit antiviral responses that are activated by IFN-α.
One of the antiviral proteins whose transcription is activated by IFN-α is PKR.
When a cell is infected with a virus, IFN-α induces expression of PKR to
bolster that which is already in the cell. The PKR responds to double stranded
RNA and binds to it. This causes it to autophosphorylate to become active
and then it phosphorylates translation initiation factor eIF-2α. The
phosphorylation eIF-2α causes protein synthesis to stop which in turn
prevents viral proteins from being synthesized. Because VP35 can inhibit
PKR, it is able to keep the cellular machinery working so that more virions
can be produced.
The viruses also target the dendritic cells early in the infection which inhibits
the adaptive immune response. Dentritic cells play a role in the immune
system by capturing and presenting foreign antigens to activate the T-cell
response. T-cells recognize the specific foreign antigens that have been
presented to them by the dendritic cells and target the cells infected with
them for destruction. The virus’ infection of dendritic cells is believed to lead
to the apoptosis of T-cells. When filoviruses attack the cells involved in
adaptive immunity, they prevent the development of response targeted
towards the specific virus.
After taking out the immune system, filoviruses are able to infect a broad
range of cells. Typically high titers of virus have been found in the liver,
spleen, and lungs of victims. Infection of the cells in these organs leads to
necrosis and eventual organ failure. Contributing to the shutdown of the
organs is the deposition of large amounts of fiber in the tissues by
macrophages. Heart failure is also commonly found in victims due to high
levels of nitric oxide in the blood. Nitric oxide helps maintain pressure in the
arteries, but overproduction can be stimulated by presence of a virus or
bacterium. It is possible that hepatocytes release large amounts of nitric oxide
in response to the filovirus infection, and consequently cause a shutdown of
the cardiovascular system.
The hemorrhaging that gives the syndrome its name results from destruction
of the vascular system. The vascular endothelium separates the blood from
tissue in the human body and its barrier is maintained by cell to cell
adherence. When this barrier weakens, blood leaks into the tissue causing
hypotension and edema. Filoviruses are known to destroy the integrity of the
vascular endothelium such that the blood vessels become permeabilized and
bleeding out occurs. Some suggest this may be due to the glycoprotein (GP)
of the virion targeting reticuloendothelial cells in order to damage vascular
structure. It has been shown the GP exhibits preferential binding to the
endothelium. Exposure of endothelial cells to the GP protein causes cell
rounding and detachment from the monolayer and a downregulation of β1integrin, which is important to cell adherence. The GP is also thought to direct
the virion to monocytes and macrophages to stimulate a cytokine induced
inflammatory response that further weakens the vascular endothelial barrier.
Currently there are no vaccines nor any standard post-infection therapy
established for the filoviruses though the use of anticoagulants has offered
some supportive care. Recently a post-exposure vaccine, similar to that of
rabies, has been developed using a live, attenuated recombinant vesicular
stomatitis virus expressing the glycoprotein of Ebola Zaire. The basis for the
vaccine is that the Ebola GP that normally protrude from the surface of the
virion should induce an immune response. The vaccine was tested on
Rhesus macaques and was given to them within half an hour of infection with
Ebola Zaire. Surprisingly, 50% of monkeys survived the infection without
major symptoms and developed IgM and IgG antibodies to the virus.
Two preventative types of vaccines are also being developed, one of which
consists of naked DNA of Ebola Zaire GP or nucleoprotein (NP), and the
other consists of an Adenovirus vector expressing both GP and NP from
Ebola Zaire and Ebola Sudan. The first vaccine uses a plasmid with a
cytomegalovirus promoter that is recognized by host polymerases to express
the GP and NP. Tests showed that this plasmid expression of either GP or NP
induced both a cell-mediated and a humoral immune response and offered
some protection from the virus in mice. The second vaccine utilizes the high
expression levels of Adenovirus type 5 and the fact that Adenovirus targets
dendritic cells which are involved in antigen presentation. Testing showed this
vaccine protected 100% of cynomolgus macaques from infection with Ebola
when given a high enough dose.
RNA (RNAi) silencing is a method of controlling cell differentiation and
developmental processes with the use of microRNAs in Eukaryotic cells. It
has also been shown to have an antiviral role in plants, nematodes and
insects. When a cell is infected with a virus, a double-stranded RNA that is
specific to the virus is produced. The double-stranded RNA is processed into
siRNAs which then become a part of the RNA-induced silencing complex.
This complex then targets viral mRNAs for destruction. The siRNA produced
in the cell also activates the secretion of type I IFNs. To evade this antiviral
activity, some viruses encode RNA silencing suppressors (RSS). Thus far,
RSSs have been shown to suppress RNAi by either binding up siRNAs or
inhibition of Dicer activity. Ebola virus protein VP35 has been shown to act as
an RSS, allowing for high replication of the virus and evasion of an antiviral
response.
In order for the filoviruses to begin the assembly and budding process, the
viral nucleocapsid (NC) is formed from genomic RNA, L polymerase protein,
VP35, NP and VP30 with NP being at the core. It is thought that NP first
forms itself into helical tubes, and VP35 and VP24 work with these tubes to
form the main NC structure. Then the NC is transported to the plasma
membrane via a cooperation between VP40 and microtubules. GP
colocalizes with VP40 to lipid rafts on the membrane, though it has been
shown that lipid rafts are not essential to the budding process. However, it is
hypothesized that the lipid rafts may aid in the curving of the membrane that
occurs during budding. The filoviruses have two modes of budding.
Filamentous virus like particles lacking the NC are able to bud and release
from the cell vertically, while mature virions bud horizontally. Interestingly,
VP40 can bud by itself as a filamentous virus-like particle.
References
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