Specific Viral Families: Poxviridae

Poxviridae

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The poxviruses, another group of enveloped, linear dsDNA viruses, are the largest and most complex of all viruses. There are terminal repeated sequences and inverted repeats at both ends. They are widely distributed in nature. Nearly every animal species can be infected by a form of poxvirus. The human poxviruses (orthopoxviruses) are large, enveloped, brick-shaped viruses. These viruses multiply in specialized portions of the host cell cytoplasm called viroplasm, where they can cause skin lesions typical of smallpox and coxpox. Other poxviruses, such as monkeypox, can infect humans who have close contact with infected animals.

Important properties

    · Large family of viruses

    · Large enough just to be seen under a light microscope.

    · Unique oval-shaped

    · Antigenically very complex

    · Unlike the influenza virus, poxviruses can remain stable for hours in the air. It could be due to the fact that it is an enveloped virus.

Smallpox virus is an example of a poxvirus.

Transmission

    · Human is the only reservoir for smallpox

    · Respiratory secretions (aerosolized air droplets) and saliva and bodily fluids from infected person are sources of transmission

Clinical Features

    · ~12 days incubation

    · Initially influenza like symptoms

    · Characteristic pustules

    · Always associated with skin lesions and scarring of skins

    · Neurological damage

    · Blindness

    · Death

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Two varieties of smallpox are:

1 Variola Major

a. 25%-30% fatalities.

  2 Variola Minor

a. Less than 1% death

Vaccination

In 1796, Edward Jenner developed a smallpox vaccine using cowpox. He used cowpox from milkmaids and inoculated it into an 8 year old boy James Phipps. When challenged with smallpox, he recovered quickly and gained immunity to smallpox.

Eradication of Smallpox

    · The variola virus has no reservoir except man

    · It only has a single, stable serotype, which is the key success of vaccination.

    · It causes only acute infection from which the patient dies, or

    · Obtains lifelong immunity to it as

    · Variola virus is an effective immunogen (sure to gain immunity from it)

    · Vaccination programs

    · Certified by WHO in 1980 to be defeated and eradicated off the face of the earth.

Laboratory Diagnosis

    · Diagnosis was made either by growing the virus in cell culture or chick embryos or by detecting viral antigens in vesicular fluid by immunofluorescence.

Control

    · Vaccination

    · Incineration of all infected material.

    · However, vaccination no longer required after 1980, when smallpox was eradicated.

    · Smallpox remains as one of the deadliest virus around—it is still kept in certain laboratories. There have been concerns that smallpox may be used in biowarfare and bioterrorism.

Iceberg Concept of Infection

Infection occurs

Specific Viral Families: Flaviviridae

Flaviviridae

The flaviviruses are enveloped, polyhedral, (+) sense RNA viruses that are transmitted by mosquitoes and ticks. The viruses produce a variety of encephalitides or fevers in humans. The yellow fever virus is a flavivirus that cause a haemorrhagic fever—in which blood vessels in the skin, mucous membranes and internal organs bleed uncontrollably. Hepatitis C infection is also caused by a flavivirus.

Important Properties

    · Spherical enveloped virion, 40-50nm

    · Inner core protein C

    · Membrane/matrix protein M

    · Envelope with glycoprotein peplomers (E)

    · Single linear 11kb positive sense ssRNA—infectious mRNA

    · Polyadenylated tail and 5’ cap

    · Cytoplasmic replication

    · Polyprotein from genomic RNA cleaved

    · 3 structural proteins

    · Several non-structural proteins

Dengue Virus

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  · Most important arbovirus presently.

    · Transmitted through the bite of a female Aedes aegypti mosquito (the vector).

    · 4 distinct serotypes based on neutralisation test.

    · DEN-1, DEN-2, DEN-3 and DEN-4

    · DEN-2 shows greatest antigenic and genotypic distance from the others.

    · Protective immunity after infection homotypic.

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Dengue Infectious Cycle

Classic Dengue begins suddenly with an influenza syndrome consisting of fever, malaise, cough, and headache. Severe pains in muscles and joints occur. Enlarged lymph nodes, a masculopapular rash, and leukopenia are common. After a week or so, the symptoms regress but weakness may persist. Although unpleasant, this typical form of dengue is rarely fatal and has few sequelae.

Dengue Haemorrhagic Fever (DHF) is a much more severe disease, with a fatality rate that approaches 10%. The initial picture is the same as classic dengue, but then shock and haemorrhage, especially into the gastrointestinal tract and skin, develop. Dengue haemorrhagic fever occurs particularly in southern Asia, whereas the classical form is found in tropical areas worldwide.

Haemorrhagic Shock Syndrome (DSS) is due to the production of large amounts of cross-reacting antibody at the time of a second dengue infection. The pathogenesis is as dollows: The patient recovers from classic dengue caused by one of the four serotypes, and antibody against that serotype is produced. When the patient is infected with another serotype of dengue virus, an anamnestic, heterotypic response occurs, and large amounts of cross-reacting antibody to the first serotype are produced.

Pathogenesis of DHF/DSS

  1. Virulent strain theory

a. Some strains are more virulent than others

b. Molecular studies show variations in sequences amongst different strains within serotypes

c. Early evidence pointed to DEN-2

  2. Antibody enhancement

a. Main theory for DHF/DSS

b. Main cell target of DEN: Monocytes and macrophages

c. Most cases of DHF/DSS had prior infection or infants below 1 year had maternal Ab.

d. Monkey experiments showed similar enhancement.

Control of Dengue Virus

    · No antiviral therapy or vaccine for dengue is available.

    · Outbreaks are controlled by using insecticides and draining stagnant water that serves as the breeding ground for mosquitoes.

    · Personal protection includes using mosquito repellent and wearing clothing that covers the entire body.

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Yellow Fever Virus

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  · Type species of the genus flavivirus

    · Tropical disease in Latin America and Africa

    · Transmitted through the bite of a female Aedes aegypti mosquito (the vector).

    · Incubation period: 3-6 days

    · Acute yellow fever—symptoms: Viremia, headache, malaise, nausea, muscle ache, flushing of head and neck, conjuntival infection, strawberry tongue.

    · Remission after acute yellow fever manifests as severe yellow fever—symptoms: fever, vomiting, abdominal pain, prostration.

    · Symptoms progress to involve the liver, kidneys and heart: e.g. jaundice.

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Strawberry Tongue

Control of Yellow Fever Virus

    · Mosquito control (similar to dengue)

    · Immunnization with the vaccine containing live, attenuated yellow fever virus. Travelers to and residents of endemic areas should be immunized.

    · Protection lasts for up to 10 years; booster shots to be taken after that.

    · Because it is a live vaccine, it should not be given to immunocompromised people or to pregnant women.

West Nile Virus

  · Originated in Uganda

· Discovered in 1937

Symptoms

    · Usually mild to no symptoms

o Fever

o Headache, body aches

o Skin rash

o Swollen lymph glands

    · Severe Symptoms (occurring mainly in persons above 50 years old)

o Crossing blood-brain-barrier

o Encephalitis

o Meningitis

Transmission

    · Wild birds are the main reservoir of this virus

    · Vector is the mosquito, especially the Culex species

    · Humans are dead-end hosts

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West Nile Virus Transmission Cycle

Control

    · No vaccine is available for west nile virus

    · Mosquito control

Specific Viral Families: Retroviridae

Retroviridae

The retroviruses are enveloped viruses that have two complete copies of (+) sense RNA. They also contain the enzyme reverse transcriptase which uses the viral RNA to form a complementary strand of DNA, which is then replicated to form a dsDNA. For the genetic information to be transcribed and translated, this DNA migrates to the host cell nucleus and become incorporated into chromosomes of host cells. Such integrated viral DNA is known as a provirus.

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Retroviruses cause tumours and leukemia in humans. The human retroviruses invade immune defense cells called T lymphocytes and are referred to as Human T cell Leukemia Viruses (HTLV). Both HTLV-1 and HTLV-2 are associated with malignancies (leukemia and other tumours), whereas the human immunodeficiency virus (HIV) causes acquired immune deficiency syndrome (AIDS).

Important Properties

    · Spherical enveloped virion, 80-100nm

    · Ribonucleoprotein in central nucleoid (concentric or truncated cone in lentivurses) within icosahedral capsid.

    · Envelope with glycoprotein peplomers

    · 2 copies of linear plus sense ssRNA each 7-10kb

    · 3’ polyadenylated tail and 5’ cap

    · Reverse transcriptase

    · Formation of long terminal repeats before prpovirus DNA inserted into host genome

    · Genome consists of gag, pol, env genes some regulatory genes, some oncogenes.

Human Immunodeficiency Virus

HIV is one of the human T-cell lymphotropic retroviruses. HIV preferentially infects and kills helper (CD4) T lymphocytes, resulting in a loss of cell mediated immunity and a high probability that the host will develop opportunistic infections. Other cells (e.g. macrophages and monocytes) that have CD4 proteins on their surfaces can be infected also.

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Genes and Proteins of the Human Immunodeficiency Virus

Gene	Proteins Encoded by Gene	Function of Proteins
I. Structural Genes Found in All Retroviruses
gag	p24, p7 p17	Nucleocapsid Matrix
pol	Reverse Transciptase Protease Integrase	Transcribes RNA genome into DNA Cleaves precursor polypeptide Integrates viral DNA into host cell DNA
env	gp120 gp41	Attachment to CD4 protein Fusion with host cell
II. Regulatory Genes Found in Human Immunodeficiency Virus that are Required for Replication
tat	Tat	Activation of transcription of viral genes
rev	Rev	Transport of late mRNAs from nucleus to cytoplasm
III. Regulatory Genes Found in Human Immunodeficiency Virus that are NOT Required for Replication (accessory genes)
nef	Nef	Decreases CD4 proteins and class I MHC proteins on surface of infected cells; induces death of uninfected cytotoxic T cells; important for pathogenesis by SIV
vif	Vif	Stabilizes newly synthesized viral DNA and transports core through cytoplasm
vpr	Vpr	Transports viral core from cytoplasm into nucleus in non-dividng cells
vpu	Vpu	Enhances virion release from cell

Replicative Cycle of HIV

The replication of HIV follows the typical retroviral cycle.

  1. Inital step in the entry of HIV into the cell is the binding of the virion gp120 envelope protein to the CD4 protein on the cell surface.

  2. The virion gp120 protein then interacts with a second protein on the cell surface, one of the chemokine receptors (CXCR4 and CCR5). Individuals with mutations in the gene encoding CCR5 have protection from infection with HIV.

  3. The virion gp42 protein mediates fusion of the viral envelope with the cell membrane, and the virion enters the cell.

  4. After uncoating, the virion RNA-dependent DNA polymerase transcribes the genome RNA into double-stranded DNA, which integrates into the host cell DNA, mediated by integrase.

  5. The viral DNA can integrate at different sites in the host cell DNA, and multiple copies of viral DNA can integrate.

  6. Viral mRNA is transcribed from the proviral DNA by host cell ENA polymerase and translated into several large polyproteins.

  7. Precursor polyproteins can be assembled into nucleocapsid.

  8. As immature virions buds from the cell membrane, the polyproteins are cleaved by the viral protease. This results in the formation of mature, infectious virions.

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Transmission

    · Transfer of infected blood.

    · Sexual contact (horizontal transfer)

    · Perinatal transmission from infected mother to neonate: Either across the placenta, at birth, or via breast milk. (vertical transfer)

Pathogenesis

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  · Primary Infection

o Acute stage

o Flu-like symptoms

o Fever

o Skin rash

o Swollen skin nodes

o Due to virulence factors such as rate of replication, propensity to mutate and cytopathogenicity.

o Also due to host resistance mechanisms such as the suppression by CD8 T suppressor cells and the presence of cytotoxic T-lymphocytes.

    · Asymptomatic Stage

o No apparent disease

o Fall in CD4 T lymphocytes

o Possible signs:

§ Fatigue

§ Depression

§ Weight loss

§ Memory disorders

    · Symptomatic Stage

o AIDS-related complex

§ Diseases not considered definitive of AIDS

§ May be attributed to HIV infection

§ Indicative of defect in cell-mediate immunity

o AIDS

§ Opportunistic infections as a result of fall in CD4 T lymphocytes. (e.g. Kaposi’s Sarcoma)

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Treatment

    · Highly active antiretroviral therapy (HAART)

o Consists of two nucleoside inhibitors (zidovudine and lamivudine) and a protease inhibitor (indinavir)

o Effective in prolonging life, improving the quality of life, and reducing viral load but does not cure the chronic HIV infection i.e. a latent infection of CD4+ cells continues indefinitely.

o Such combination therapy able to combat resistance—which arise due to rapid mutations due to inefficiency of reverse transcriptase.

    · Non-specific therapeutic management

    · Immunomodulation

o Enhancement of immune system through treatment with interleukin-2 (still under study)

    · No vaccine for human use is available.

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Prevention

Taking measures to avoid exposure to the virus:

    · Using condoms

    · Not sharing needles

    · Discarding donated blood that is contaminated with HIV

Specific Viral Families: Orthomyxoviridae

Orthomyxoviridae

The orthomyxoviruses are medium-sized, enveloped, (-)-sense RNA viruses that vary in shape from spherical to helical. Their genome is segmented into eight pieces. Orthomyxoviruses have an affinity for mucus. The influenza A and B viruses in this viral family are of particular medical importance to humans since they cause disease in humans.

Influenza Viruses

The influenza A virus is an orthomyxovirus that infects humans, birds, swine and horses. It causes pandemics of influenza (~10-20 years) and major outbreaks of influenza (virtually every year in various countries). The influenza B virus, on the other hand, appears to be specific to humans only. It causes major outbreaks of influenza, which occur less often as those caused by the influenza A virus.

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Diagram of the Influenza virus

Important Properties

    · Typically spherical (100nm in diameter).

    · Composed of a single stranded (-) sense RNA genome in eight segments, a helical nucleocapsid, and an outer lipoprotein envelope.

    · 3 polymerase polypeptides with each segment.

    · 5’ and 3’ end of all segments are highly conserved.

    · Virion contains RNA-dependent RNA polymerase, which transcribes the (-) sense RNA to mRNA. Thus, the genome is not infectious.

    · Viral envelope is covered with two different types of spikes—haemagglutinin (HA) and neuraminidase (NA).

    · The ratio of HA:NA is 5:1.

Haemagglutinin functions at the beginning of infection, whereas the neuraminidase functions at the end.

Haemagglutinin

    · Binds to the cell surface receptor (neuraminic acid, sialic acid) to initiate infection.

    · Target of neutralizing antibody.

Neuraminidase

    · Cleaves neuraminic acid (sialic acid) to release progeny virus from the infected cell.

    · Degrades the protective layer of mucus in the respiratory tract, thus enhancing the ability of the virus to infect the respiratory epithelium.

Pathogenesis

    · After the virus has been inhaled, the neuraminidase degrades the protective mucus layer, allowing the virus to gain access to the cells of the upper and lower respiratory tract.

    · The infection is limited primarily to this area because the HA receptors have a specific affinity for the epithelial cells of the respiratory tract.

    · Despite systemic symptoms, viremia rarely occurs.

    · The systemic symptoms are due to cytokines circulating in the blood.

    · There is necrosis of the superficial layers of the respiratory epithelium.

    · Immunity rests mainly upon secretory IgA in the respiratory tract.

    · IgG is also produced and confers protection against future infections.

    · Innate resistance also plays a role in immunity—a mucus blanket and cilia helps to trap and expel the influenza virus out of the respiratory tract.

    · Cytotoxic T cells also play a protective role.

    · Immunocompromised patients especially prone to secondary infection such as pneumonia.

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Clinical Features

    · After an incubation period 24-48 hours, fever, myalgias, headache, sore throat, and cough develop suddenly.

    · Severe myalgias (muscle pains) coupled with respiratory tract symptoms are typical of influenza.

    · Vomiting and diarrhoea are rare.

    · Symptoms usually resolve spontaneously in 4-7 days.

Differentiating Between the Common Cold and Flu

Laboratory Diagnosis

    · Nasal/throat washings or swabs; sputum.

    · Virus culture in MDCK cell line or chick embryo.

    · Direct ELISA for Haemagglutinin

    · PCR assay

Transmission and Epidemiology

    · Virus is transmitted by airborne respiratory droplets.

    · Ability of influenza A virus to cause epidemics is dependent on antigenic changes in haemagglutinin and neuraminidase.

    · Influenza infections are found all year round.

    · In the northern hemisphere, influenza occurs primarily in the winter months.

    · In the southern hemisphere, influenza occurs primarily in the winter months of June through August.

    · Few serotypes circulating simultaneously.

    · Epidemics and pandemics occur when the antigenicity of the virus has changed sufficiently that the pre-existing immunity of many people is no longer effective.

    · The antigenicity of influenza B virus also varies but not as dramatically or as often.

Antigenic Drift vs. Antigenic Shift

Antigenic Drift	Antigenic Shift
Minor change in genome.	Major change in genome.
Occurs in both Influenza A and B viruses.	Occurs in only Influenza A virus.
Point mutation in the genetic code of surface antigens.	Gene reassortment, where entire segments of RNA are exchanged, each one of which codes for a single protein (e.g. haemagglutinin).
Results in a new strain.	Results in a new subtype.
Happens all the time.	Happens occasionally.
Responsible for epidemics.	Responsible for pandemics.

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Strain Naming Convention

Strains of influenza are named as follows:

(Influenza Type)/(Animal species (omitted if human)/(Place of Isolation)/(Number of Isolates)/(Year of First Isolation)(H and N subtypes)

Example: A/Texas/36/91/(H1N1)

Control and Treatment

    · The main mode of prevention is the vaccine, which consists of killed influenza A and B viruses. It is rendered useless by antigenic shift/drift—therefore, the vaccine is usually reformulated every year to contain the current antigenic strains.

    · Antiviral drugs are used to treat influenza. This includes receptor analogues, transcriptase inhibitors, reverse transcriptase inhibitors, protease inhibitors and neuraminidase inhibitors etc.

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