Friday, January 9, 2009

Picornaviridae and Orthomyxoviridae


Introduction to Picornaviridae
‘Pico’ is very small and it is also known as Pico PNA viruses.

Picornavirus is one of the ‘oldest’ known viruses, dating back to 1400 B.C., when a temple record tells of poliovirus in ancient Egypt. Picornavirus is one of the most diverse viruses with over 200 sereotypes, causing infections such as Polio, Hepatitis A, and the common flu. Foot-and-mouth virus was one of the first viruses to be recognised in 1898 by Loeffler and Frosch, which cause infections in livestock.

Classification of Picornaviridae
Base on physical properties, the particle density and pH-sensitivity, and serological relatedness, more recently based on nucleotide sequence. The most recent revision of virus taxonomy has recognized nine genera within the family:

Picornaviruses are classed under Baltimore's viral classification system as group IV viruses as they contain a single stranded, positive sense RNA genome of between 7.2 and 9.0 kb in length. Like most positive sense RNA genomes, the genetic material alone is infectious; although substantially less virulent than if contained within the viral particle, the RNA can have increased infectivity when transfected into cells. The genome itself is the same sense as mammalian mRNA, being read 5’ to 3’. Unlike mammalian mRNA Picornaviruses do not have a 5’ CAP but a virally encoded protein known as VPg, however like mammalian mRNA the genome does have a poly A tail at the 3’ end. There is an un-translated region (UTR) at both ends of the Picornavirus genome. The 5’ UTR is longer, being around 600-1200 BP in length, compared to that of the 3’ UTR, which is around 50-100bp. It is thought that the 5’ UTR is important in translation and the 3’ in negative strand synthesis; however the 5’ end may also have a role to play in virulence of the virus. The rest of the genome encodes structural proteins at the 5’ end and non-structural proteins at the 3’ end in a single polyprotein.

The whole of replication occurs within the host cell cytoplasm and infection can even happen in cells that do not contain a nucleus (known as enucleated cells) and those treated with actinomycin D (this antibiotic would inhibit viral replication if this occurred in the nucleus.)

Genome structure of Picornaviridae

The picornavirus genome consists of a single molecule of linear, postitive(+)-sense, single-strand RNA and is non-segmented. The complete genome is nucleotides long. The 5'-terminus of the genome has a long untranslated region 600-1200 bases in length, which is important in translation, virulence, and possibly encapsidation. There is a shorter untranslated region (50-100 bases in length) on the 3'-terminus, which is important in (-) strand-synthesis. The 5'-terminus untranslated region also has a "clover leaf" secondary structure known as the Internal Ribosome Entry Site (IRES), which distinguishes picornaviruses from other RNA viruses; this structure is important in translation and replication. The 5'-terminus is modified by a covalently-attached VPg protein (which takes the place of a cap), while the 3'-terminus is modified by polyadenylation.
Virion structure of a Picornaviridae
Picornavirus virions consist of a non-enveloped, icosahedrally symmetric capsid. The capsid consists of 12 capsomers and has a diameter of 27-30 nm, which makes it one of the smallest of all viruses (thus the name "picornavirus"). The genome is tightly packed into the capsid. The capsid has four unique proteins: VP1, 2, 3, and 4.
Reproduction cycle of Picornaviridae
Replication of picornaviridae
Using different cellular receptors (depending on the picornavirus), a picornavirus virion attaches to a host cell. Uncoating occurs, and the virus' RNA is released into the cytoplasm of the host cell through a membrane channel. Virus replication occurs entirely in the cytoplasm. The host cell's transcription processes are shut off to a degree that varies with different picornaviruses, while the IRES helps to make sure the virus' transcription is left untouched. Replication occurs. RNA is packaged into preformed capsids. Release of the virus occurs when cell lysis occurs (with the exception of Hepatits A, which is non-lytic and thus creates a more persistent infection).

Viral ecology and pathology
The mode of transmission, ecology, and pathology of picornaviruses vary greatly between the different genera. Interestingly, the genetics of virulence phenotypes of picornaviruses is poorly understood.

Picornaviruses do not have virulence genes per se, but the design of the capsid and how it interacts with the virus receptor expressed on the host cell surface, specific sequences within the nontranslated regions of the viral genome, as well as coding sequences that result in different protein sequences may all have a part in determining the virulence phenotype.

Transmission and effects of picornaviridae
Transmission of picornaviridae

Effect of picornavirus on human
Introduction to orthomyxoviridae
‘Ortho’ is the Greek for straight and ‘Myxa’ is the Greek word mucus. It has a common name, Avian influenza, bird flu, fowl plague.

These viruses cause influenza, an acute respiratory disease with prominent systemic symptoms. Pneumonia may develop as a complication and may be fatal, particularly in elderly persons with underlying chronic disease. Type A viruses cause periodic worldwide epidemics (pandemics); both types A and B cause recurring regional and local epidemics. Influenza epidemics have been recorded throughout history. In temperate climates, the epidemics typically occur in the winter and cause considerable morbidity in all age groups. An epidemic with associated mortality has occurred in most of the past 100 years. The worst of these was the 1918 pandemic, which caused about 20 million deaths worldwide and about 500,000 deaths in the United States.

Classification of orthomyxoviridae
The orthomyxoviridae are a family of RNA viruses that includes the 5 generas, Influenzavirus A, Influenzavirus B, Influenzavirus C, Isavirus and Thogotovirus. Influenza A, B and C causes influenza in vertebrates, like for example, birds, humans and other mammals. Isavirus infect salmon while thogotoviruses infect vertebrates and invertebrates such as mosquitoes and sea lice.
Animal influenza virus
Genome structure of orthomyxoviridae

The genome of the orthomyxovirus consists of six to eight segments of linear, negative-sense, single-stranded RNA. The complete genome is 10000-14600 nucleotides long. Segment 1 is fully sequenced and the complete sequence is nucleotides long. Segment 2 only has an estimate of the sequence, although it is sequenced and is almost the same length as segment 1. Segment 3 is also sequenced, but estimated and the complete sequence is around 2200-2300 nucleotides long. Segment 4 has been completely sequenced and it is 1700-1800 nucleotides long. Segment 5 has been sequenced, but only estimated, and is 300-1900 nucleotides long. Segment 6 has been sequenced, but only estimated, and is 1400-1500 nucleotides long. Segment 7 has been sequenced, but estimated and the complete sequence is 800-1100 nucleotides long.

The genome has terminally redundant sequences and they are repeated at both ends. The nucleotide sequences at the 3'-terminus are identical. The 5'-terminal sequence has conserved regions and repeats complementary to the 3'-terminus; terminal repeats at the 5'-end are 11-14 nucleotides long. The 3'-terminus has conserved nucleotide sequences; is 11-13 nucleotides long; in the genera of same family. The sequence has conserved regions in all RNA species or some RNA segments. The multipartite genome is encapsidated with each segment in a separate nucleocapsid, and the nucleocapsids are surrounded by one envelope. Each virion contains defective interfering copies.

Viririon structure of orthomyxoviridae
Virions of an orthomyxovirus consist of an envelope, a matrix protein, a nucleoprotein complex, a nucleocapsid, and a polymerase complex. The virus capsid is enveloped. Lipids are present and located in the envelope. Virions are composed of 18-37% lipids by weight. The composition of viral lipids and host cell membranes are similar. The lipids are modified cellular lipids and are derived from plasma membranes. Proteins of host derived membranes have been modified during post-translational processes. Host-derived membranes contain viral proteins in place of host proteins.

The virions are spherical to pleomorphic and filamentous forms occur. They are 80-120 nm in diameter and 200-300(-3000) nm long. The surface projections are densely dispersed distinctive hemagglitinin-esterase (HEF) spikes, or spaced widely apart hemagglutinin (HA) spikes. Clusters of neuramidase (NA) irregularly inerpose the major glycoprotein in a ratio of HA to NA about 4-5 to 1.

Human influenza viruses
(Envelope contains rigid "spikes" of haemagglutinin and neuraminidase which form a characteristic halo of projections around negatively stained virus particles.)
There are about 500 spikes evenly dispersed or clustered and are covering the surface comprising hemagglutinin, or neuraminidase, or esterase-esterase. The surface projections are composed of one type of protein or different types of proteins and are 10-14 nm long and 4-6 nm in diameter. The nucleocapsid is elongated with helical symmetry and is segmented with loops at one end. The segments have different sized classes with clear predominate lengths with a length of 50-130 nm (in differnent class sizes) and a width of 9-15 nm.

Reproduction cycle of orthomyxoviridae

Orthomyxovirus replication takes about 6 hours to replicate and kills the host cell. The viruses attach to permissive cells via the hemagglutinin subunit, which binds to cell membrane glycolipids or glycoproteins containing N-acetylneuraminic acid, the receptor for virus adsorption. The virus is then engulfed by pinocytosis into endosomes. Acid environment of the endosome causes the virus envelope to fuse with the plasma membrane of the endosome, uncoating the nucleocapsid and releasing it into the cytoplasm.

A transmembrane protein derived from the matrix gene (M2) forms an ion channel for protons to enter the virion and destabilize protein binding allowing the nucleocapsid to be transported to the nucleus, where genome is transcribed by viral enzymes to yield viral mRNA. Unlike replication of other RNA viruses, orthomyxovirus replication is dependent on the presence of active host cell DNA. The virus scavenges cap sequences from the nascent mRNA generated in the nucleus by transcription of the host DNA and attaches them to its own mRNA. These cap sequences allow the viral mRNA to be transported to the cytoplasm, where it is translated by host ribosomes. The nucleocapsid is assembled in the nucleus.

Virions acquire an envelope and undergo maturation as they bud through the host cell membrane. During budding, the viral envelope hemagglutinin is subjected to proteolytic cleavage by host enzymes. This process is necessary for the released particles to be infectious. Newly synthesized virions have surface glycoproteins which contain N acetylneuraminic acid as a part of their carbohydrate structure, and thus are vulnerable to self-agglutination by the hemagglutinin. One major function of the viral neuraminidase is to remove these residues.

Gene Reassortment
The influenza virus genome is segmented, and because of that, genetic reassortment can occur when a host cell is infected simultaneously with viruses of two different parent strains. If a cell is infected with two strains of type A influenza, for example, some of the progeny virions will contain a mixture of genome segments from the two strains. This process of genetic reassortment probably accounts for the periodic appearance of the novel type A strains that cause influenza pandemics

Viral ecology and pathology
Influenza virus is transmitted from person to person primarily in droplets released by sneezing and coughing. Some of the inhaled virus lands in the lower respiratory tract, and the primary site of the disease is the tracheobronchial tree, although the nasopharynx is also involved. The neuraminidase of the viral envelope may act on the N-acetylneuraminic acid residues in mucus to produce liquefaction. This liquified mucus may help spread the virus through the respiratory tract in concert with the mucociliary transport. The superficial mucosa suffers cellular destrcution and desquamation because of the infection of the mucosal cells. Nonproductive cough, sore throat and nasal discharge are some sysmptoms that result from the endema and mononuclear cell infiltration of the involved areas. The cough may be persistent but the most prominent symptoms of influenza are systemic-- fever, muscle aches and general prostration. These systemic symptoms are not caused directly by the virus because viremia is rare. A possible cause is circulating interferon, as administration of theraupetic interferon causes systemic symptoms resembling those of influenza.

The evidence shows that the extent of virus-induced cellular destruction is the prime factor determining the occurence, severity and duration of clinical illness. It is possible to recover virus from respiratory secretions for 3 to 8 days in an uncomplicated case. At times of maximal illness, peak quantities of 104 to 107 infectious units/ml are detected. The titer begis to drop in concert with the progressive abatement of disease after 1 to 4 days of peak shedding

The infection may extensively involve the alveoli, particularly in patients with underlying heart or lung disease. This may result in interstitial pneumonia, sometimes with marked accumulation of lung hemorrhage and endema. Pure viral pneumonia of this type is a severe illness with a high mortality. Virus titers in secretions are high, and viral shedding is prolonged. However, in most cases, bacteria are the causative agent of pneumonia associated with influenza. Examples include pneumococci, staphylococci, and Gram-negative bacteria. The preceding viral infection damages the normal defenses of the lung, setting the stage for the bacteria to invade and cause disease.

Host defences
Immune mechanisms are responsibe for recovery from influenza have not been clearly delineated. Several mechanisms probably act in concert. Interferon appears in respiratory secretions shortly after viral titers reach their peak level, and may play a role in the subsequent reduction in viral shedding. Antibody usually is not detected in serum or secretions until later in recovery or during convalescence; nevertheless, local antibody appears responsible for the final clearing of virus from secretions. T cells and antibody-dependent cell-mediated cytotoxicity also participate in clearing the infection.

Antibody is the primary defense in immunity to reinfection. IgG antibody, which predominates in lower respiratory secretions, appears to be the most important. The IgG in these secretions is derived from the serum, which accounts for the close correlation between serum antibody titer and resistance to influenza. IgA antibody, which predominates in upper respiratory secretions, is less persistent than IgG but also contributes to immunity.

Only antibody directed against the hemagglutinin is able to prevent infection. A sufficient titer of anti-hemagglutinin antibody will prevent infection. Lower titers of anti-hemagglutinin antibody lessen the severity of infection. Anti-hemagglutinin antibody administered after an infection is under way reduces the number of infectious units released from infected cells, presumably because the divalent antibody aggregates many virions into a single infectious unit. Antibody directed against the neuraminidase also reduces the number of infectious units (and thus the intensity of disease), presumably by impairing the action of neuraminidase against N-acetylneuraminic acid residues in the virion envelope and thus promoting virus aggregation. Antibody directed against nucleoprotein has no effect on virus infectivity or on the course of disease.

Immunity to an influenza virus strain lasts for many years. Recurrent cases of influenza are caused primarily by antigenically different strains.


Electron micrograph of influenza virus particles
Diagnosis of influenza is suggested by the clinical picture of sudden onset of fever, malaise, headache, marked muscle aches, sore throat, nonproductive cough, and coryza. When a syndrome resembling influenza occurs in the winter in an adult (the etiologies of illnesses of this type are more complex in children), an influenza virus is a likely cause. If an epidemic of febrile respiratory disease is known to be under way in the community, the diagnosis is yet more likely. Definitive diagnosis, however, relies on detecting either the virus or a significant rise in antibody titer between acute phase and convalescent-phase sera.

A rapid specific diagnosis of influenza may be obtained by demonstrating viral antigens in cells obtained from the nasopharynx in immunostaining tests such as immunofluorescence or in enzyme immunoassays (ELISA) employing respiratory secretions. Influenza virus is usually isolated from respiratory secretions by being grown in tissue cultures or chick embryos. Virus growth in tissue cultures is detected by testing for hemadsorption: red cells are added to the culture and adhere to virus budding from infected cells. If the culture tests positive, serologic tests with specific antisera may be used to identify the virus. In the chick embryo culture method, fluid from the amniotic or allantoic cavity of chick embryos is tested for the presence of newly formed viral hemagglutinin; the virus in positive fluids is then identified by hemagglutination inhibition tests with specific antisera. Finally, a rise in serum antibody titer between acute-phase and convalescent-phase sera can be identified by various tests, of which complement fixation, hemagglutination inhibition, and immunodiffusion (using specific viral antigens) are the most common. None of these techniques will identify all infections.

Prevention and treatment
Inactivated influenza virus vaccines have been used for about 40 years to prevent influenza. The viruses for the vaccine are grown in chick embryos, inactivated by formalin, purified to some extent, and adjusted to a dosage known to elicit an antibody response in most individuals. A given vaccine contains the strains of types A and B viruses that are judged most likely to produce epidemics during the following winter. Vaccine is administered parenterally in the fall; one or two doses are required, depending on the immune experience of the population with related antigens. Protection against illness has varied from 50 to 90 percent in civilian populations and from 70 to 90 percent in military populations. Local and systemic reactions to the vaccine are minor and occur in the first day or two after vaccination. During the national swine flu immunization of 1976 in the United States, an increased risk of developing Guillain-Barre syndrome accompanied vaccination; however, this correlation has not been detected since. Annual use of inactivated influenza virus vaccine is currently recommended in the United States for persons at risk of developing pneumonia from the disease and for their close associates. Live attenuated vaccines are being developed as alternatives to inactivated vaccine.

The synthetic drugs amantadine and rimantadine hydrochloride effectively prevent infection and illness caused by type A, but not by type B, viruses. The drugs interfere with virus uncoating and transport by blocking the transmembrane M2 ion channel (see multiplication). Drugs prevent about 50 percent of infections and about 67 percent of illnesses under natural conditions. When administered for 10 days to household contacts of a person with influenza, drugs protect up to 80 percent of the persons from illness. Side effects are greater for amantadine and limited primarily to the central nervous system.

Amantadine and rimantadine are the only specific antiviral treatments available for influenza. As in the case of prophylaxis, they are effective only against type A virus. When administration is started early in the course of illness, drugs hasten the disappearance of fever and other symptoms. Emergence of viral resistance can occur during treatment.


1 comment:

Sheldon said...

These viruses can be detected by using ELISA kits. Many hospitals have been using this. These are great help in the field of medicine and technology.