Viral Genetics

).Salwa Hassan Teama )M.D

Contents
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Viruses Structure Virus Replication Laboratory Diagnosis Genetic Changes in Viruses Viruses and Cancer

Viruses

Viruses are particles composed of an internal core containing
either DNA or RNA (but not both) covered by a protective protein coat. Some viruses have an outer lipoprotein membrane, called an envelope, external to the coat.

Viruses are not considered free-living, since they cannot

reproduce outside of a living cell; they have evolved to transmit their genetic information from one cell to another for the purpose of replication.

Viruses, can affect virtually all life forms, including humans,
animals, plants, fungi, and bacteria.

Viruses

Viral infections are the most cause of human disease, and are
responsible for at least 60% of the illness.

Viral infections may be broadly classified into those in which the

immune response eliminates the virus from the body )e.g. influenza and polio viruses) and those in which the virus is able to persist despite the host immune response. Viruses may persist as a latent infection, with or without intermittent replication )e.g. herpes simplex virus), or as a chronic infection )e.g. HIV or HCV virus).

Examples of common human diseases caused by viruses include the common cold, influenza, chickenpox, diarrhea and cold sores. Serious diseases such as Ebola, AIDS, avian influenza and SARS are caused by viruses.

Viruses

Viruses often damage or kill the cells that they infect, causing disease in infected organisms. A few viruses stimulate cells to grow uncontrollably and produce cancers. Antibiotics have no effect on viruses, but antiviral drugs have been developed to treat life-threatening infections. Vaccines that produce lifelong immunity can prevent viral infections.

Virus Structure

Structure
Size & Shape  Viral Nucleic Acids  Viral Capsid and Symmetry  Viral Proteins

Size and Shape
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Viruses vary in size 20 to 300 nm in diameter. Most viruses appear as spheres or rods in the electron microscope. In addition to these forms, bacterial viruses can have very complex shapes such as T4 phage. The shape of virus particles is determined by the arrangement of the repeating subunits that form the protein coat (capsid) of the virus.

Viral Capsid and Symmetry

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The protein coat (capsid), made up of subunits called capsomers. The structure of the nucleic acid genome and the capsid protein is called the nucleocapsid. The arrangement of capsomers gives the virus structure its geometric symmetry. The viral nucleocapsids have two forms of symmetry either helical or icosahedral.

Helical

• Icosahedral.

Viral Nucleic Acids
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The viral nucleic acid (genome) is located internally and can either single or double sranded DNA or single or double stranded RNA. The nucleic acid can be either linear or circular. The DNA is always a single molecule; the RNA exist either as a single molecule or in several pieces. e.g. both influenza virus and rotavirus have a segmented RNA genome.

Viral Protein
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Viral proteins serve several important functions, The outer capsid proteins protect the genetic material and mediate the attachment of the virus to specific receptors on the host cell surface. It is also important antigens that induce neutralizing antibody and activate cytotoxic T cells to kill virus infected cells. Viruses also have internal viral proteins, some of which are DNA or RNA polymerases. Some viruses produce antigenic variants of their surface proteins that allow the viruses to evade our host defenses.

Viral Envelope

Lipoprotein membrane composed of lipid derived from the host cell membrane and protein that is virus specific. The presence of an envelope confers instability on the virus. Enveloped viruses are more sensitive to heat, drying, detergent and lipid solvent such as alcohol and ether than are nonenveloped. Enveloped viruses are transmitted by direct contact via blood and body fluids, whereas naked viruses can survive longer in the environment and can be transmitted by indirect means such as feco-oral route.

Atypical Virus Like Agents

Defective viruses are composed of viral nucleic acid and proteins but cannot replicate without a helper virus which provide the missing function. Pseudivirions contain host cell DNA instead of viral DNA within the capsid. Viroids consist of single molecule of circular RNA without a protein coat or envelope. Prios are infectious particles that composed of proteins and contain no detectable nucleic acid.

Virus life cycle

The life cycle of viruses differs greatly between species but there are six basic stages

Virus Life Cycle
2. 3. 4. 5. 6. 7.

Attachment is a specific binding between viral capsid proteins and specific receptors on the host cellular surface, Penetration , viruses enter the host cell through receptor mediated endocytosis or membrane fusion, Uncoating , the viral capsid is degraded by viral enzymes or host enzymes thus releasing the viral genomic nucleic acid, Replication involves synthesis of viral messenger RNA ) mRNA( for viruses except positive sense RNA viruses, Viral protein synthesis and assembly of viral proteins and viral genome replication, Viruses are released from the host cell by lysis. Enveloped viruses (e.g., HIV) typically are released from the host cell by budding.

Virus Life Cycle

Classification of Medically Important Viruses
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I -dsDNA viruses : e.g. Adenoviruses, Herpesviruses, Poxviruses II- ssDNA viruses(+) : sense DNA e.g. Parvoviruses III- dsRNA viruses : e.g. Reoviruses IV- ssRNA viruses :(+) sense RNA e.g. Picornaviruses, Togaviruses V- ssRNA viruses : (-) sense RNA e.g. Orthomyxoviruses, Rhabdoviruses VI- ssRNA-RT viruses :(+) sense RNA with DNA intermediate in life-cycle e.g. Retroviruses VII- dsDNA-RT viruses : e.g. Hepadnaviruses

Laboratory Diagnosis
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Identification of the virus in cell culture; Microscopic identification directly in the specimen; Serologic procedures to detect a rise in antibody titer or the presence of Ig M antibody; Detection of viral antigen in blood or body fluids; Detection of viral nucleic acids in blood or patients cells.

Identification of The Virus in Cell Culture
The presence of a virus in a patient's specimen can be detected by cytopathic effect in cell culture but CPE is not specific. A specific identification of the virus usually involves an antibody based test as fluorescent antibody, complement fixation or ELISA.

Microscopic identification directly in the specimen
Electron microscopy is the most common method used to study the morphology of viruses.
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Inclusion bodies, formed by aggregates of many virus particles, can be seen in either the nucleus or cytoplasm of infected cells. Multinucleated giant cells are formed by several viruses e.g. herpes, respiratory syncytical virus and measles virus. Fluorescent antibody staining of cells obtained from the patients or of the cells infected in culture can provide a rapid specific diagnosis. Electron microscopy is not often used in clinical diagnosis but is useful in the diagnosis of certain virus e.g Ebola (characteristic appearance and are dangerous to grow in culture).

Serologic Procedures
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The presence of Ig M antibody can be used to diagnose current infection. The presence of Ig G antibody cannot be used to diagnose current infection. Rise in antibody titer that is 4 fold or greater in the convalescent serum sample compared to the acute sample can be used to make a diagnosis.

Detection of Viral Antigen & Nucleic Acids

The presence of hepatitis B surface antigen is commonly used in diagnosis. The presence of vial DNA or RNA is increasingly becoming the gold standard in viral diagnosis. Molecular diagnostic procedures have been available since 1970s,when researchers first began using cloned DNA probe to detect viral nucleic acid. The new molecular diagnostic methods predicted that nucleic acid tests would rapidly replace traditional virus detection methods.

Molecular Diagnostic Methods
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The goal is in the detection of non culturable agents such as human papilloma virus, human parvovirus, Detecting viruses difficult to culture, including enteric adenovirus, some coxsackie viruses, Detecting viruses that are dangerous to culture such as HIV, Detecting viruses that are present in low numbers, for example, HIV in antibody negative patients or CMV in transplanted organs. Important when a tiny volume of specimen is available (forensic samples or intra-ocular fluid specimens). Allow laboratory to predict antiviral drug susceptibilities and to detect infections when viable virus cannot be obtained (latent viral infection or viruses that are present in immune complexes). May also used to differentiate antigenically similar viruses such as adenovirus types 40 and 41 and to detect viral genotypes that are associated with human cancers (human papilloma virus).

Viral Genetics

Viruses grow rapidly, there are usually a large number of progeny virions per cell. There is, therefore, more chance of mutations occurring over a short time period. The nature of the viral genome (RNA or DNA; segmented or nonsegmented) plays an important role in the genetics of the virus.
Viruses undergo genetic change by several mechanisms :

Genetic drift where individual bases in the DNA or RNA mutate to other bases. Antigenic shift is where there is a major change in the genome of the virus. This occurs as a result of recombination.

Mutants
Spontaneous mutations
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These arise naturally during viral replication: e.g. due to errors by the genome-replicating polymerase or a a result of the incorporation of tautomeric forms of the bases. DNA viruses tend to more genetically stable than RNA viruses. There are error correction mechanisms in the host cell for DNA repair, but probably not for RNA.

Induced mutation by physical or chemical means
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Chemical agents : e.g. nitrous acid,………… Physical: agents such as UV light or X-rays,..

Recombination

Process of intermolecular exchange, of chromosomes combining genetic information from different sources, typically two genomes of a given species. This kind of break/join recombination is common in DNA viruses or those RNA viruses which have a DNA phase (retroviruses).

Reassortment
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Reassortment, mixing of the genetic material of two similar viruses that are infecting the same cell. Reassortment of a virus has a segmented genome. Human viruses characterized with segmented genomes are RNA viruses e.g. orthomyxoviruses, reoviruses, arenaviruses, bunya viruses

generating novel reassortants and has also been useful in laboratory experiments.  Reassortment is a non-classical kind of recombination.

Reassortment may play an important role in nature in

Viruses and Cancer

A limited number of DNA viruses are associated with certain types

of human neoplasia, while a variety of RNA viruses, or retroviruses, cause tumors in animals…. The main viruses associated with human cancers are human papillomavirus, hepatitis B and hepatitis C virus , Epstein-Barr virus, and human T-lymphotropic virus,……….. Viruses seem able to cause cancer in three ways Presence of the viral DNA may disrupt normal host DNA function. Viral proteins needed for virus replication may also affect normal host gene regulation. The virus may serve as a vector for oncogene insertion.

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References& Online Further Reading
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Basic Virology, In. Review of Medical Microbiology and Immunology. Eds) Warren Levinson).ISBN. 0-07110438-0 Daniel H. Farkas. DNA Simplified: The Hitchhiker's Guide to DNA. Washington, DC: AACC Press, 1996, ISBN 0-915274-84-1. William B. Coleman,Gregory J. Tsongalis: Molecular Diagnostics: For the Clinical Laboratorian: ISBN 1588293564... Daniel P. Stites,Abba T. Terr. Basic Human Immunology: ISBN. 0838505430 Danny L.Wiedbruk and Ann M Drevon. Nuclaic acid detection methods . 1995 by academic press. Diamandis E.1993. The role of clinical chemistry in molecular diagnostics. Clin. Chem. News 19,4. Kulski JK and Norval M 1985 Nucleic acid probes in diagnosis of viral diseases of man. Brief review . Arch. Virology.83,3-15. Low JB 1986. Clinical application of gene probes in human genetic disease, malignancy ,and infectious diseas.Clin.Chem.Acta.157,1-32. Ray CG 1979. An infectious disease viewpoint of diagnostic virology. In diagnosis of viral infection.pp241248.University Park Press Baltimore. Tenover FC1988 Diagnostic DNA probes for infectious disease.Clin.Microbiol.Res.1,82-101 http://biology.about.com/library/weekly/aa110900a.htm http://www-micro.msb.le.ac.uk/3035/HBV.html http://cancerweb.ncl.ac.uk/cgi-bin/omd?tautomerism http://pathmicro.med.sc.edu/lecture/RETRO.HTM

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