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Home , History of virology

1 I. GENERAL Introduction Virology is the study of and viral . This book focuses on broader aspects of virology. These are summarized as follows: • Virology as a scientific discipline (History). • General properties and structure of viruses. • Taxonomic nature of viruses • Way, in which it is cultured, purified and assayed. • Viruses of bacteria • Viruses of plants. • Viruses of animals. • Viruses of algae • Viruses of fungi • Viruses of insects • Prophylactic measures of viral infections in plants and animals

1.GENERAL PROPERTIES OF VIRUSES Viruses are submicroscopic, unique group of infectious agents and are obligate intracellular parasites. They contain single type of . Viruses exist as two phases. They are intracellular and extracellular viruses. Size ranges from 20 -14,000 nm in length. Complete particle is called as virion.

Definition Luria and Darnell in 1968 defined viruses as entities whose genome is an element of nucleic acid either DNA or RNA. It reproduces inside of living cells and use their synthetic machinery to direct the synthesis of extracellular infectious viral particles are called virion which contain the viral genome and transfer it to the other cell. Novel properties of viruses ▪ Viruses are ultramicroscopic in size ranging from 10 nm to 450 nm. Smallest viruses are little larger than ribosomes. Largest viruses are like smallest bacteria. ▪ Viruses possess very compact structure. ▪ Viruses are acellular in nature. Because they do not independently fulfill the characteristic of life. They require host cell for their replication and protein synthesis. ▪ Viruses are crystalizable in nature. ▪ Basic structure of virus consists of protein and nucleic acid. ▪ Capsid is made up of repeating unit called capsomer. It protects nucleic acid. Nucleic acid can be either DNA or RNA but not both. ▪ Nucleic acid types include single stranded RNA, double standard RNA, double standard DNA and single stranded DNA. ▪ Molecules on cell surface impart high specificity for host cell. Spike proteins of enveloped virus and capsid protein of non-enveloped virus are responsible for antigenicity and pathogenicity of viruses. ▪ Viruses lack machinery for synthesizing proteins. They are obligate intracellular parasites of bacteria, protozoa, fungi, algae, plants and animals. ▪ All the viruses are infectious in nature. ▪ Viral genome is replicated within an appropriate host cell and directs the synthesis of viral components by host cellular system. ▪ Progeny viruses are formed by assembly. Assembled viruses are disassembled during next progeny. ▪ Viruses do not have any cell organelles like chloroplast, ribosomes, mitochondria etc. ▪ A virus contains single genome only (either DNA or RNA). Viruses differ from living cells by 1. Their simple, acellular organization. 2. Absence of both nucleic acid (RNA and DNA) in same virion. 3. Their inability reproduces independently.. 2 4. There is no cell division as like prokaryotes and eukaryotes.

2. HISTORY OF VIROLOGY The term virus is derived from Latin word. It means “poison”. To understand what virus are and where they belongs in the hierarchy of biological system. It is strengthened by looking at the history. The real history of scientific evaluation starts from 18th century. The prehistoric period starts from about 4000 BC itself. Ancient people were aware of virus and they also carried research into causes and prevention of diseases. A hieroglyph from Memphis. Memphis is the capital of ancient Egypt drawn in about 3700 BC depicts a temple priests, which shows clinical sign of paralytic poliomyelitis. Smallpox is endemic in China during 1000BC. Pharoh Ramresv was succumbed to smallpox in 1196 BC. He was mummified in Cairo museum. The pustule lesion is similar to recent patients. Historical events of Virology 1976 Edward He is an English physician collects pustule fluid from Sarah Nelmsa milkmaid. He Jenner suffered from cowpox and injected into a teen age boy James Phippes. Boy develops fever and head ache and recovered fully. Then Jenner inoculated Phippes with live smallpox pustule. He did not develop small pox. This process is called vaccination. Thus he made the importance of 1798 Edward Small pox vaccine was developed Jenner 1885 Developed vaccine, described the term virus (Poison) and vaccination (to honor Jenner) 1886 John Buist He described elementary bodies of small pox from skin lessions. 1886 Adolf Mayer He demonstrated that sap of infected plant transmit disease to healthy plants.

1892 Dmitri He was a Russian botanist. He provided evidence for virus causation of Mosaic Iwanowski Disease of Tobacco(TMV). He presented a paper to the St. Petersburg Academy (1864-1920) of Science which showed that extracts from diseased tobacco plants could transmit disease to other plants. This was generally recognised as the beginning of Virology. Unfortunately, Iwanowski did not fully realized the significance of these results. 1898 Martinus He proved that a virus particle causes the Tobacco mosaic disease and described Beijerinck Contagium Virus Fluidum or living infectious fluid. He extended Iwanowski's results on and developed modern idea of the virus. Loeffler and He discovered Foot and Mouth Disease virus (FMD). In 1898, Fredrich Loeffler Frosch and Paul Frosch showed that a similar agent was responsible for foot-and-mouth disease in cattle. These agents caused disease in animals as well as plants. 1900 He demonstrated that is caused by a virus. It was spreaded by mosquitoes. 1903 Remlinger He discovered Rabies virus 1903 A.Negri He observed inclusion bodies from rabies infected human brain cells called Negri bodies

3 1908 Karl He proved that poliomyelitis is caused by Virus Landsteiner and Erwin Popper 1911 Franas He discovered causative agent for chicken cancer and Named Rous Sarcoma Peyton Rous Virus. Awarded Nobel prize 1966 1915 Frederic He discovered viruses infecting bacteria. Twort (in 1915) and Felix d'Herelle (in 1917) were the first to recognize viruses which infect bacteria. 1917 Felix He discovered the viruses of bacteria and coined the term (eaters D.Herelle of bacteria) 1932 Used mice as the host for virus 1933 R.E.Shope He identified viruses causing cancer. 1935 Wendel He crystallized TMV and showed its infectious nature. Awarded Nobel prize in stanley 1946 1938 Max Theiler He developed live attenuated vaccine against yellow fever. He was awarded Nobel prize in 1951. This discovery eventually enabled Max Theiler (1937) to propagate the virus in chick embryos and successfully produced an attenuated vaccine. 1939 Emory Ellis They established concept of one-step virus growth cycle. They were awarded and Max Nobel prize in 1969 Delbruck 1940 Helmut He used to take first viral pictures. Ruska 1941 George Hirst He demonstrated that virus agglutinates RBC. 1942 JJ .Bittner He discovered mammalian RNA tumour virus (mouse mammary tumour virus). 1945 S.Luria and He demonstrated mutation. A.Hershy 1949 John ender, They demonstrated the growth of virus using human culture. They T.Weller, were the Nobel prize winners of the year 1954. F.Robbins 1950 A.Lwoff, Discovered lysogenic bacteriophage in Bacillus megaterium. Coined the term L.Siminovuc prophage. They were awarded the Nobel prize in 1965 h and N.Kjeldgaar d 1951 Theiler He developed yellow fever vaccine

4 1952 Renato He demonstrated that also for plaques in a similar way of phages. He was Dulbecco awarded Nobel prize in 1975. 1953 NP.Rowe He discovered Adenovirus. 1955 FL Schaffer Polio virus was crystallized 1957 Fraenkel- He demonstrated that when mixtures of TMV protein and RNA were incubated Contrat and together viral particles formed spontaneously. Discovered RNA as a genetic material Williams of TMV 1963 Baruch He discovered HBV. He was awarded Nobel prize in 1976. Blumberg 1966 Edgar and He discovered pathways of Macromolecular assembly. Wood 1967 Theoder He discovered viroids Diener 1970 Howard He discovered in Retro virus Temin and 1972 Paul berg Created first rDNA in SV40 virus contain Lambda phage and galactose operon. 1977 Frederick Sequenced θx174 genome and demonstated it had 5375 nucleotides Sanger 1979 Small pox was officially declared as eradicated disease. Declared by WHO. 1981 Yorio He isolated T cell Leukaemia virus (HTLV) Hinuma 1981 DG Kleid FMD vaccine prepared by rDNA technology. 1982 Panicali and virus used as vaccine. Paoletti 1983 Luc Discovered HIV, causative agent for AIDS. Montagnier and R.Gallow

1986 Wang HDV identified First HBV vaccine produced by genetic engineering method and approved for human use. 1989 HCV was identified. 1990 First human gene therapy was approved. 1994 Yuan Chang Human Herpes Virus 8 was identified, responsible for Koposis sarcoma. and P.Moore 1995 Chicken pox vaccine was approver for US use .1997 S.Prusiner Discovered Prions. 1999 Nucleotide sequence of large known virus genome Paramecium bursaria chlorella virus was completed 2002 Corona virus was identified as a causative agent of SARS.

3.CLASSIFICATION OF VIRUSES

In 1962, Lowff R.W.Horne and P.Tournier classified all viruses based on Linnaean hierarchical system. Although a subsequently formed International committee on taxonomy of Viruses (ICTV) accepted many principles but didn’t adapt this system in Toto. Four important properties of current viral classification are 5 ▪ Nature of nucleic acid ▪ Symmetry of capsid ▪ Presence or absence of envelope ▪ Dimension of the virion Latest classification of ICTV (1995) include 21 animal virus family, 6000 viruses, totally 75 family of viruses, 9 sub family , 164 genera. ICTV system also includes prions, viroids and satellites. Order of presentation in classification ▪ ds DNA viruses ▪ ss DNA viruses ▪ RNA and DNA reverse transcribing viruses ▪ ds RNA viruses ▪ Negative strand ss RNA viruses ▪ Positive strand ss RNA viruses ▪ Subviral agents Viral properties used in taxonomy • Virion morphology- size, shape, capsid symmetry, envelope availability. • Genome structure • Sensitivity to physical and chemical agents • Viral constituents • Antigenic properties • Replication strategy • Host range • Mode of transmission • Pathogenicity Nomenclature This involves the use of specific suffixes. Genera - Virus Subfamily - Virinae Family - Viridae Order - Virales 6 Example Order Family Subfamily Genus Species Paramyxo Respiroviru Sendaivirus virinae s

Rubulavirus Measlesvirus

Morbilivirus

Pneumo virinae Pneumoviru HumanRespiratory s Syncytialvirus Metapneum Turkeyrhinotracheit

o virus is

Mononegavirales virus

Baltimore Classification David Baltimore proposed a classification system for animal viruses based on genetic system. Main properties adapted for this classification are nucleic acid and its polarity. mRNA is defined as positive strand because it contains immediately translatable information’s. DNA strand of equalant polarity is designated as + strand. The DNA and RNA complement of + strand are designated as – strand. Numbering system is adapted in system. Nucleic acid and its polarity Baltimore classification +/-DNA -I +DNA -II +/-RNA -III +RNA -RNA -IV -RNA -V -RNA +RNA -VI Baltimore system Animal viral group I Papovaviridae II III IV Calciviridae Picornaviridae Togaviridae V Bunyaviridae Paramyxoviridae Arenaviridae VI Retroviridae

7 LHT CLASSIFICATION OF VIRUS Various groups of animal viruses are available. They causes diseases to animals. Based on various features of viruses the following classification was done. The following characters are adopted to this classifications. They are, 1. Morphological features. 2. Based on Physiochemical properties. 3. Based on the genomic nature. 4. Based on the presence of protein. 5. Specificity of host. 6. Organ specificity. 7. Based on the site of multiplication within host cell. 8. Transcriptional variation. 9. Serological variation. 10. Mode of transmission. 1) MORPHOLOGICAL FEATURES i. Virion size ❖ Virions range in size from about 10 nm to 300 or 400 nm in diameter. ❖ Smallest viruses are little larger than ribosomes. Eg. Parvo virus-18 to 22nm. ❖ Largest viruses are about the same size as the smallest bacteria .Eg: Poxviruses upto 400nm. Adenovirus 60 nm to 90 nm. Parvovirus 18 nm . Reovirus 75 nm to 80 nm. . 22 nm to 30 nm. Calcivirus 35 nm to 40 nm. . 80 nm to 100 nm. ii. Virion Shape Viruses are classified in to following types based on shape. They are a. Icosahedral in shape : Adenovirus b. Helical shape : Baculovirus. c. Bullet shaped : Rhabdovirus. d. Brick shaped : Poxvirus. e. Complex shaped : Poxvirus f. Cylindrical shaped : Baculovirus. iii. Based on the envelope Many viruses have an envelope , an outer membranous layer surounding the nucleocapsid. They are enveloped viruses and non enveloped viruses. Enveloped viruses are sensitive to disinfectants, solvents etc; Eg. Enveloped viruses – Rhabdoviruses.

Non enveloped viruses –Adenoviruses. Herpesvirus. Poxvirus. ENVELOPED Togovirus. Rhabdovirus. Retrovirus. NONENVELOPED Papovavirus. Parvovirus. Adenovirus. 8 Picorno,calcivirus. iv. Capsid Symmetry it is based on capsid symmetry. This kind of viruses are classified into two groups, namely 1. Icosahedral – Herpesvirus. 2. Helical – Tobacco mosaic virus. v. Presence and absence of peplomers Peplomers are the protein projection on the surface of the capsidcalled spikes. Spikes may be involved in the attachment to the host cell surface. Eg. Adenovirus- spike present. Picornavirus - spike absent. 2) Based on physico chemical properties i. Molecular mass(Weight) Molecular mass means the weight of the virion. Small sized viruses are low molecular weight particle and larger viruses are high molecular mass particle. ii.Sedimentation Coeficient The sedimentation rate of the particle during centrifugation is determined three forces. They are, Centrifugal force, Buoyant force of the medium and fractional resistance to the particle movement. Eg : Poxvirus - 170S, Parvovirus - 20S iii. Solvents and detergents action In general, enveloped viruses are sensitive to solvent action. Naked viruses are resistant to solvent action. Coronaviruses - sensitive to solvents. Reoviruses - resistant to solvents. Plasmaviruses - sensitive to detergents. Adenoviruses – resistant to detergents.

3. Based on the genome nature i. Type of Nucleic acid The genome of viruses contains either DNA or RNA. It is based on the genome of viruses. They are classified as DNA viruses and RNA viruses. DNA virus - Herpes virus. RNA virus - Orthomyxo virus. ii. Strandedness of Nucleic acid On the basis of the nature of genome strand the viruses are classified as ss DNA Virus - Parvovirus ds DNA virus - Poxvirus ss RNA virus - Coronavirus ds RNA virus - Reovirus iii. Nature of genome arrangement On the basis of the arrangement and assembly of the genome viruses are classified as Linear DNA - Poxvirus Circular RNA -  phage Linear RNA – Picornavirus iv. Sense of the single stranded genome It is based on the polarity of the single stranded RNA. The viruses are classified as positive sense RNA viruses and negative sense viruses. +ve sense RNA directly act as a mRNA. + sense RNA – Picornavirus. −sense RNA − Filoviridae.

v. Number of segments of the genome Some of the RNA viruses have segmented genome. 9 Eg. Reovirus – 10 to 12segments. Birnaviridae – 2 segments. Influenzavirus – 8 segments HIV virus – 2 segments. vi. G+C content of the genome Melting point of the genome and it’s strength is assessed based on the availability of G+C content ,because G+C bases are bonded with triple hydrogen bond but A+T is double bond. Due to this G+C is resistant than A+T. 51% G+C in Adenovirus. 55- 60% G+C in Herpesvirus. vii. Size of the genome The larger sized viruses contain large sized genome. The smaller sized viruses contain small sized genome. Eg.:Poxvirus contains large sized genome of 150 x 106 daltons about 270 kb pairs. Picornaviridae contains small sized genome of 2.5 x 106 daltons about 4.5 kb pairs.

4. Based on the presence of proteins Proteins are the major part of viruses. The surface proteins are the major epitopes of virus. They differ in number, size and functional activities. Non structural proteins have special functional activities. Transcriptase, reverse transcriptase, haemaagglutinin, neuraminidase have different functional activities. Eg.:Retrovirus had surface specific binding sites gp120 and gp4. It selectively binds on CD4 receptor of host. Orthomyxo virus had surface proteins like Haemeagglutinin and Neuraminidase. They selectively bind on respiratory epithelial cell receptors (Neuraminic acid).

5. Specificity of host Some of the viruses are host specific. They affect only a specific host and cause an infection. Eg:Influenza virus ‘A’ causes infection in human only. HIV causes infection only in human. Human polio causes infection in human only. 6. Organ specificity Some viruses cause disease in special organs or special type of tissues. They are called organ specific or tissue specific viruses. Eg: Picornavirus (polio) only affects CNS. HIV:Only affect the immune system, they bind only a CD4 receptor containing T4 lymphocytes.

7. Based on the multiplication within host cell Viruses are intracellular parasites, they multiply inside the host cell. Multiplication of viruses are of two types : 1)Multiplication inside the nucleus. 2) Multiplication at cytoplasmic or other cell organelles. Eg : Poxvirus – cytoplasmic replication Adenovirus – nucleus replication 8. Transcriptional variation Transcription means synthesis of mRNA from the original genome. It is used for the synthesis of proteins and . Some viruses use host RNA polymerase to synthesize mRNA and some other viruses use their own RNA polymerase to transcription process. Eg: Picorna virushave a transcriptase used for transcription process. Reovirus have a transcriptase enzyme. Filovirus uses host RNA polymerase Paramyxovirus uses host RNA polymrase 10 9. Serological Variations Serological variation is based on the surface proteins present on the viruses. It is based on antigenic variations of the species, sub types. It is also based on antigen antibody reaction, antigen bind with specific antibody use to detect the surface marker proteins. Eg:Influenza virus ‘A’:H1 N1, H2 N2, H3 N3, Hn Nn. Influenza virus ‘B’: Bt . and are the types of serological variations among viruses. Eg:Influenza virus.

10. Mode of transmission The viruses are transmitted between the host by any one of the following methods. Eg: Air borne : Influenza virus. Blood borne : HIV, Yellow fever. Water borne : Entric virus , Polio. By contact : Poxvirus. Sexual contact : HIV , Yellow fever. Animal bite : Rabies virus.

4. Ultra structure of viruses Viruses are a unique group of infectious agents. The complete viral particle is called virion, consist of one or more molecules of DNA or RNA enclosed in a coat protein, sometimes also in other layers. Viruses exist in two phases they are intracellular and extracellular. Viruses are smaller than bacteria and larger than cell organelle. Viruses multiply within a host cell. Structure of Virus Virus morphology had been studied over the past decades. Electron microscopy, x-ray diffraction, biochemical analysis and immunological techniques are adopted to study the detailed morphology of viruses.

Virion Size Size of the virion ranges from about 10 - 300 or 400nm in diameter. Smallest virus is little larger than ribosomes whereas the poxviruses, are about the size as the smallest bacteria. Smallest virus –Parvovirus,  x 174 Largest virus- Poxviruses

Capsid The virus consists of a protein coat capsid and a nucleic acid core. Nucleic acid core of the viruses is surrounded by protein coat called capsid. Capsid protects the viral genetic material and aids in its transfer between host cells. The capsid is composed of a number of subunits known as capsomers. Capsomers are assembled and give rise to viral symmetry. There are three morphological types of . They are 1.Polyhedral/Icosahedral/Spherical symmetry. 2.Helical/Cylindrical/Rod like symmetry. 3.Complex/Binal symmetry. 1.Icosahedral or polyhedral It is a regular polyhedron with 20 equilateral triangular faces and 12 intersecting points. These capsids appear spherical when viewed at low power in the electron microscope. Icosahedral symmetry occurs more frequently. Watson and Crick shown the polyhedral capsids exhibit 3 types of symmetry. 11 Eg: Tetrahedral, Octahedral and Icosahedral. Viruses employ the Icosahedral shape because it is a most efficient way to enclose a space. Capsids are large macromolecular structures constructed from many copies of one or more or few types of protein subunits called protomers. Five or six protomers combined form a structure, which was represented as capsomers. A few genes can code for proteins that self assemble to form the capsid. Capsid is made up of pentamers and hexamers. 5 monomers conbines to from a pentamer and is called pentameric capscomer. Each capsid consists of many capsomers. Pentamers are at the verities, where as hexamers form its edges and triangular faces. Protomers join to form capsomers through neon covalent bondings. The bonds between proteins within pentamer and hexamers are stronger those between separate capsomers. Number of Capsomers:- The minimum number of capsomer is 12 followed by 32, 42. 72, 92, 162, 252, 362, 492, 642 and 812  x 174 – 12 TYNV – 32 Adenovirus – 252 Herpes virus – 162 Reovirus - 92 Polyyoma and Papilloma viruses – 72. Bond between two capsomers is weaker than between monomers. The capsids break down into capsomers during purification. In most of the viruses, pentamers and hexamers are made up of same type of monomers. Exception is Adenovirus.

2. Helical or cylindrical Symmetry It consists of monomers arranged in a helix around a single rotational axis. Eg. TMV. Helical capsids are shaped much like hollow tubes with protein wall size of the helical capsid is influenced by its protomers and nucleic acid enclosed within the capsid.

3. Complex or banal symmetry Complex viruses have capsid symmetry that is neither purely icosahedral nor helical. They may have fail and other structure or have multilayered wall surrounding nucleic acid. Complex viruses with both heads and tails are said to have binal symmetry because they possess a combination of icosahedral head and helical tail. Eg. Bacteriophage.

12 Many animal viruses and some plant viruses are bounded by an outer membranous layer, in addition to capsid are called an envelope. Envelope is about 10-15 nm in thickness. Chemically envelope is made up of carbohydrate protein and lipids. Lipids and proteins are associated to form lipoproteins. Carbohydrates and proteins associated and forms glycoproteins. It is made from host cell plasma membrane, but they vary considerably in size, morphology, complexity and nature of the membrane. Envelope viruses acquire their membrane, a lipid bi-layers with associated protein by budding through a cell membrane, the endoplasmic reticulum, golgi apparatus or nuclear membrane. Envelope proteins are generally called as glycoproteins that carry covalently linked sugar chains or oligosaccharide. Sugars are added to the proteins post translationally during transport to the cell membrane, from which progeny viruses assemble. Intra or Interchain disulfide bonds, another chemical feature of envelope protein. Covalent bonds stabilize the tertiary or quarternary structure of viral glycoproteins. Viral glycoproteins are embedded in the lipid bi-layer by a short membrane-spanning domain. It is a major antigenic determinant and mediate fusion during entry. Internal domains also available in the envelope and is generally called as surface proteins, which is essential for viral assembly. In some viruses, receptor destroying enzymes are available to promote viral release. Eg.Influenza ‘A’contains Neuraminidase, Corona virus contains Esterase. In Influenza ‘A’ virus, a proton channel (M2) proteins are available which may facilitate uncoating and transcriptase activity. Fusion protein is also available in the envelope and mediate fusion. In some viruses, additional viral structural proteins are available, termed the matrix protein. It mediates the interaction of the innermost, genome containing structure with the viral envelope. Examples: G Protein - Rhabdovirus HA,NA, M2 - OrthomyxoVirus Env - Retrovirus gH gL - Herpes virus

Envelope is a flexible, membranous structure so enveloped viruses can have various shaped and are pleomorphic. Envelope confer chemical, biological and antigenic properties or viruses. Envelopes are susceptible to lipid solvents like chloroform, ether and bile salts phospholipids of envelope includes phosphatidyl choline, phosphatidyl serine, phosphatidyl ethanolamine. Glycolipids include Sphingosine. Functions ❖ Gives structure ❖ Protects NA and Capsid ❖ It helps in adherence of virus on to receptors of host cell. ❖ Envelope confers chemical, biological and antigenic properties of viruses.

13 Spikes Some of the proteins project from the envelope and are called spikes or peplomers. It is composed of glycoproteins. Functions ❖ They adhere to the receptors of host cell and play an important role in infection. ❖ Spikes mediate host viral inferaction. Eg. Influenza virus, Rhabdhovirus, HIV etc.

Viral enzymes Enzymes catalyze a variety of biological reactions and are called biocatalysts. 1. DNA polymerase It is also known as DNA dependent DNA polymerase. It is used in polymerization of nucleic acid. Eg. Involved in T4 DNA replication 2. RNA polymerase RNA viruses code for RNA dependent RNA polymerase or DNA dependent RNA polymerase. These enzymes mediate viral specific protein synthesis. Eg. Poxvirus DNA dependent RNA dsDNA mRNA Polymerase

RNA dépendent RNA -ss RNA mRNA Polymerase

3. RNA dependent DNA polymerase This special enzyme is involved in the synthesis of DNA from RNA.. This enzyme is also called Reverse transcriptase. This enzyme is found in .

4.Terminal nucleotidyl transferase This enzyme will add Poly A or Poly U sequence at 3’ and 5’ end. Eg. Adenovirus, Herpesvirus

5. Integrase and protein kinase Integrase is involved in integration of host chromosome. Protein kinase is used for synthesis of protein. Other enzymes like hyaluronidase hydrolyze hyaluranic acid in respiratory track; Neuraminidase hydrolyses N acetyl muramic acid present in epithelial cells of respiratory track.

Effect of Physical and Chemical agents on viruses Heat: Viruses are inactivated at 50C for 30 minutes and at 100C for few seconds. 14 Cold: Viruses are stable at low temperatures and can be stores at - 40C or at 70C. Some viruses are partially inactivated at freezing temperatures. UV: It inactivates viruses. Chloroform and ether : They are known as organic solvents. They act on lipids. So enveloped viruses are easily inactivated as lipids are present in envelopes. Oxidising and reducing agents: Viruses are inactivated by chlorine, iodine, formaldehyde and H2O2. Phenols : Most viruses are resistant to phenols.

Viral nucleic acid Viruses are exceptionally flexible with respect to the nature of their genetic materials. They have all four possible nucleic acid types. ds DNA ds RNA ss DNA ss RNA

All the four types are found in animal and plant viruses. Phages contain DNA as genetic materials. Size of the genome varies greatly. The smallest genome are around 1 X 106 D.

Viral genetics Like other living beings viruses obey the laws of genetics. Properties of viruses such as virulence and antigenicity are under genetic control. Two main mechanisms for genetic modification in viruses are mutation and recombination.

Mutation It is defined as alterations in the genetic material leading to permanent hereditary changes. The frequency of mutation in viruses is about 104 to 108. Mutation occurs in every infection. Most mutations are lethal mutation may occur spontaneously or by induced process such as irradiation or 5 flurouracil.

Recombination Genetic recombination may occur when two related viruses infect a cell simultaneously. This may leads to antigenic drift (minor change) and antigenic shift (Major change). Eg: Influenza virus.

5. Replication of viruses Reproduction or replication of viruses is also called as Host-Virus interactions. Viruses are obligate intracellular parasite. It reproduce only within a host cell. Viruses have the specific efficiency towards the cells. Certain groups of viruses may infect only specific groups of cells. The viruses make use of the metabolic machinery of the host cell to undertake replication. Eg: Bacterial viruses only infect bacteria . Animal viruses only infect animal cells. Depends upon tissue tropism, replication or reproduction of viruses may be divided into several stages. They are • Adsorption of viruses • Penetration and uncoating • Replication of nucleic acid • Synthesis and assembly of viral capsids. • Release of mature virions Adsorption of virion • It is a first step in multiplication process. It occurs through random collision on plasma membrane of host cell. • Adherence is an important virulent mechanism of virus. 15 • Capsid protein in non-enveloped viruses and spike protein in enveloped viruses play a crucial role in attachment process. Eg: Haemagglutination spike of Influenza virus Capsid protein in Parvovirus. • Spike of non-enveloped virus also involved in attachment process. Eg. Adenovirus spike. • Infection of the virus depends greatly on its ability to bind the cell. Viruses have special structure (some times called Ligands) binds to the receptors on the host cell surface. • Receptor is a glycoprotein plays an important role in the tissue tropism and host specificity. • Attachment is in most cases a reversible process - if penetration does not ensue, the virus can elute from the cell surface. Examples • Influenza A virus binds sialic acid receptor of respiratory track. • Rabies virus adsorbs an Acetyl choline receptor of neurons. • Vaccinia virus attach on epidermal growth factor receptor • Rhino virus binds to intracellular adhesion molecules on the respiratory epithelial cells. • HIV virus binds CD4 receptor • HAV binds α 2 macroglobin. Penetration and uncoating • Viruses penetrate the plasma membrane and enter a host cell shortly after adsorption. The mechanism of penetration and uncoating must vary with different viruses. • Penetration of viruses involves 3 process. They are 1. Direct penetration. 2. Fusion. 3. Endocytosis. 1. Direct penetration (Translocation) Naked virus (Eg. ) undergoes conformational changes after adherence on plasma membrane and released only nucleic acid in to the cytoplasm. 2. Fusion Enveloped virus fuses with plasma membrane with the help of fusion protein or F protein. It results in entry of capsid protein into the cytoplasm. 3. Endocytosis Most of the enveloped viruses enter inside of host cells through receptor-mediated endocytosis and form coated vesicles. Immediately after entry uncoating taken place by making use of lysozyme or acidity formation within an endocytic vesicle. Replication of nucleic acid DNA viruses Most of the DNA virus genome is replicated within a host cell nucleus with few exceptions. Eg. Smallpoxvirus. Upon entry, genome enters into a nucleus and perform transcription by using a enzyme RNA polymerase. In some viruses, two stages of transcription occur. They are early transcription and late transcription. During transcription mRNA will be formed, which is transferred to cytoplasm for process. This process leads to the synthesis of protein responsible for DNA replication. DNA is replicated by using an enzyme DNA polymerase. This leads to the replication o genome. Replication and transcription in RNA viruses RNA viruses adopt four strategies for replication. The process of replication depends upon the nature of RNA. 16 1. Some RNA viruses use their RNA genome as a giant mRNA Eg. Poliovirus. These types of viruses are considered as + sense ss RNA viruses.

2. dsRNA viruses like Reoviruses carry a virus associated transcriptase and generates mRNA. RNA polymerase enzyme produces new dsRNA from +mRNA.

3. In case of negative sense ss RNA viruses, viral replicase converts the ss RNA into a double stranded RNA called the replicative form. This directs the synthesis of specific RNA genome.

4. Retrovirus perform a different pattern of replication. Ss RNA is copied into DNA RNA hybrid by making use of reverse transcriptase enzyme. Then the Ribonuclease H degrades +RNA strand to leave – DNA. After the synthesis of – DNA, the reverse transcriptase copies this strand to produce a double stranded DNA. It is called proviral DNA, which can direct the synthesis of mRNA and multiple copies of new + RNA virion genome.

Site of replication vary depends up on the type of viruses Virus Site of Replication Adenovirus Nucleus Hepadnovirus Cytoplasm Poxvirus " Parvovirus " Papovavirus " Orthomyxovirus Nucleus Panamyxo virus Cytoplasm Picornavirus " Retrovirus "

Synthesis and assembly of capsid Late genes of virus is responsible for the synthesis of structural proteins (capsid and spike protein). Viruses are assembled after the complete synthesis of structural and functional protein. Assembly takes 17 place in anyone of the adjacent places of plasma membrane, nuclear membrane, endoplasmic reticulum or golgi apparatus Virus Assembly Site Adenovirus Nuclear Hepadnavirus Cytoplasm Poxvirus Cytoplasm Orthomyxovirus Nucleus Rhabdovirus Cytoplasm

Virion Release It differs between naked and enveloped viruses. There are three mechanisms available to release viruses from host cells. These are cell lysis or cytopathic effect, budding and cell degeneration. Most of the naked viruses are released by lysing the host cells and results in cytopathic effect. Enveloped viruses may receive envelope from golgi apparatus, endoplasmic reticulum or plasma membrane. If the virus receives envelop from plasma membrane. It releases by budding. Others may be released by cell lysis. Viruses such as Parvoviruses accumulate within the host cells and are released only after the death of the host cell, which follows the degeneration of host cell. Actin filament of host cell can aid in virion release.

Effect of viruses on host cell 1. Inhibits host DNA, RNA and protein synthesis 2. Lysosomes may be damaged resulting in cell lysis 3. Intracellular structures called inclusion bodies are formed. 4. Produce chromosomal disruptions. 5. May produce transformed cells (Cancer cells). 18 Virus and host interaction () may leads to following types of infection. Some type of interaction leads to cancer. • Productive infection – complete replication of infections virion and release of virus. • Abortive infection – synthesis of without production of infection virion. • Semi permissive infection – complete replication with low yield of infection virion. • Malignant transformation – associated with integration of viral DNA and differential viral and cellular gene expression. • Viral latency – persistence of viral genome in the host cell.