The Fascinating World of Viruses

1. Chapter 1: Viruses

2. Introduction to Viruses • “Virus” originates from Latin word “poison”.Term was originally used by Pasteur to describe infectious agent for rabies. First virus discovered was tobacco mosaic disease virus (TMV) in 1890s. In 1930s: TMV was isolated and purified. Electron microscope was used to observe viruses. • Viruses - The Boundary of Life (www.dellpassovoy.com/Viruses.ppt - Mirip ) At the boundary of life, between the macromolecules (which are not alive) and the prokaryotic cells (which are), lie the viruses and bacteriophages. In isolation, viruses and bacteriophages show none of the expected signs of life. They do not do any of the things we normally associate with life. Strictly speaking, they should not be considered "living" organisms at all. However, they are more complex than a lifeless collection of macromolecules and they do show one of the most important signs of life: the ability to reproduce at a fantastic rate but only in a host cell.

3. Characteristics of all viruses • Acellular infectious agents. A virus is a non-cellular particle made up of genetic material and protein that can invade living cells. (www.worldofteaching.com/powerpoints/biology/viruses.ppt - Mirip ) • Possess either DNA or RNA, never both Replication is directed by viral nucleic acid within a cell • A virus is an obligate intracellular parasite containing genetic material surrounded by protein (http://www.slideworld.org/viewslides.aspx/Introduction-to-Virology-ppt-67425) • Lack genes and enzymes necessary for energy production • Depend on host cell ribosomes, enzymes, and nutrients for protein production • Incapable of independent metabolisms

4. Viruses are Smaller Than Most Cells

5. Components of mature viruses (virions): Capsid: Protein coat made up of many protein subunits capsomeres). Capsomere proteins may be identical or different. Genetic Material: Either RNA or DNA, not both Nucleocapsid = Capsid + Genetic Material Additionally some viruses have an Envelope(consists of proteins, glycoproteins, and host lipids.Derived from host membranes. Naked viruses lack envelopes.

6. Viruses Have Either DNA or RNA Inside a Protein Capsid (Nucleocapsid) Influenza Virus Bacteriophage

7. Viruses are classified by the following characteristics: • Type of genetic material • Capsid shape • Number of capsomeres • Size of capsid • Presence or absence of envelope • Host infected • Type of disease produced • Target cell • Immunological properties

8. Types of viral genetic material: Genetic material may be single stranded or double stranded: • Single stranded DNA (ssDNA): • Parvoviruses • Double stranded DNA (dsDNA): • Herpesviruses • Adenoviruses • Poxviruses • Hepadnaviruses* (Partially double stranded) • Single stranded RNA (ssRNA): May be plus (+) or minus (-) sense: • Picornaviruses (+) • Retroviruses (+) • Rhabdoviruses (-) • Double stranded RNA (dsRNA): • Reoviruses

9. Capsid morphology: • Polyhedral: Many-sides. Most common shape is icosahedron, with 20 triangular faces and 12 corners. • Poliovirus • Herpesvirus • Complex viruses: Unusual shapes • Helical: Ribbon-like protein forms a spiral around the nucleic acid. May be rigid or flexible. • Tobacco mosaic virus • Ebola virus • Bacteriophages have tail fibers, sheath, and a plate attached to capsid. • Poxviruses have several coats around the nucleic acid.

10. Active Virus • ATTACHMENT: A specific virus attaches to the surface of a specific cell • INVADE: The nucleic acid (DNA or RNA) of the virus is injected into the cell. (Note : ~ Penetration etc.) • COPY: The viral nucleic acid takes control of the cell an begins to make new virus particles. (Note : ~ Replication etc.) • RELEASE: The cell bursts open, hundreds of new virus particles are released from the cell. These virus particles go on to infect other cells. (Note : ~ Lysis ---- although not always lysis/rupturing host cell)

11. Life Cycle of Animal Viruses • Attachment or adsorption: Virus binds to specific receptors (proteins or glycoproteins) on the cell surface. • Penetration: Virus enters cell through one of the following processes: • Direct fusion with cell membrane • Endocytosis through a clathrin coated pit • Uncoating: Separation of viral nucleic acid from protein capsid. Lysosomal, cytoplasmic, or viral enzymes may be involved.

12. Life Cycle -Animal Viruses (Continued) 4. Synthetic Phase: (Involves several processes) • Synthesis of viral proteins in cytoplasm • Replication of viral genome: • DNA viruses typically replicate in nucleus • RNA viruses replicate in cytoplasm • Assembly of progeny virus particles The synthetic stage can be divided in two periods: • Early period: Synthesis of proteins required for replication of viral genetic material. • Late period: Nucleic acid replication and synthesis of capsid and envelope proteins

13. Life Cycle-Animal Viruses (Continued) 5. Release of progeny virions: There are two main mechanisms of release A. Lysis of cells: Naked viruses and pox viruses leave cell by rupturing the cell membrane. Usually results in death of the host cell. Example: Poliovirus B. Budding: Enveloped viruses incorporate viral proteins in specific areas of a membrane and bud through the membrane. Envelope contains host lipids and carbohydrates. Host cell does not necessarily die. Example: Human Immunodeficiency Virus

14. Life Cycle of a DNA Virus

15. Life Cycle of Bacteriophages Lytic Cycle: Cell bursts at end of cycle 1. Attachment or adsorption: Virus tail binds to specific receptors on the cell surface. 2. Penetration: Virus injects genetic material (DNA) into cell. Tail releases lysozyme, capsid remains outside. 3. Biosynthesis: Viral proteins and nucleic acids are made. 4. Maturation: Bacteriophage capsids and DNA are assembled into complete virions. 5. Release: Bacteriophage virions are released from the cell. Plasma membrane breaks open and cell lyses. Burst time: Time from attachment to release of new virion (20-40 minutes). Burst size: Number of new phage particles that emerge from a single cell (50-200).

16. Lytic Cycle of Bacteriophage

17. Lytic Cycle of Bacteriophage

18. Life Cycle of Bacteriophages Lysogenic Cycle 1. Attachment and Penetration: Virus tail binds to specific receptors on the cell surface and injects genetic material (DNA) into cell. 2. Circularization: Phage DNA circularizes and enters either lytic or lysogenic cycle. Lysogenic Cycle 3. Integration: Phage DNA integrates with bacterial chromosome and becomes a prophage. Prophage remains latent. 4. Excision: Prophage DNA is removed due to a stimulus (e.g.: chemicals, UV radiation) and initiates a lytic cycle.

19. Lysogenic versus Lytic Cycles of Bacteriophage

20. Retroviruses Convert RNA into DNA via Reverse Transcriptase

21. How does our body respond to viruses?www.scripps.edu/community/lessonplans/Gibson%201.ppt - Amerika Serikat Immunobiology, 5th ed. Janeway

22. HUMAN IMUNODEFICIENCY VIRUS (HIV) • Ex.of a Retrovirus is HIV www.dellpassovoy.com/Viruses.ppt - Mirip • A typical, "minimal" retrovirus consists of: • an outer envelope which was derived from the plasma membrane of its host • copies of an envelope protein embedded in the lipid bilayer of its envelope • a capsid; a protein shell • two molecules of RNA and • molecules of the enzyme reverse transcriptase www.worldofteaching.com

23. Retrovirus www.worldofteaching.com

24. Viruses are host specific – a protein on the surface of the virus has a shape that matches a molecule in the plasma membrane of its host, allowing the virus to lock onto the host cell. HIV doesn’t target just any cell, it goes right for the cells that want to kill it. “Helper" T cells are HIV's primary target. These cells help direct the immune system's response to various pathogens. • www.dellpassovoy.com/Viruses.ppt - Mirip

25. The HIV in humans can only bind and fuse to the white blood cells, so therefore it lowers the CD4+ count and in turn, the efficiency of the immune system. Also, since the virus can only infect the white blood cells, it cannot be transmitted by touch or by inhalation. • Once a white blood cell is infected, the human immune system tries to build up its stamina in order to fight the disease; however, it is during this first period that the HIV virus is able to replicate extensively. After this first period of time, the immune system brings the virus under control and levels off the dropping number of CD4 cells. This time period is when humans may live for long amounts of time without any outside effects or illness. • Although the number of CD4 cells is now stable, the HIV virus continues to multiply & infect. This is called the asymptomatic stage of infection. By this time, the amount of CD4+ T cells being destroyed almost equal to the amount of viruses destroyed. This "full-scale war" between the two has roughly 10 billion virus particles being made and destroyed and roughly 1 billion CD4+ T cells being killed and replaced each day. • This dynamic equilibruim lasts for about 10 years or so until the enitre immune system collapses, and the beginning of the onset of AIDS. The cause of death can range from Tuberculosis to mold infections that affect the brain, liver, or intestines. (e.g.Cryptococcus meningitis, Toxoplasmosis brain abscesses, gastroenteritis etc.) to cancers caused by viruses.

26. HIV undermines the body's ability to protect against disease by depleting T cells thus destroying the immune system.The virus can infect 10 billion cells a day, yet only 1.8 billion can be replaced daily. • After many years of a constant battle, the body has insufficient numbers of T-Cells to mount an immune response against infections. At the point when the body is unable to fight off infections, a person is said to have the disease AIDS. • It is not the virus or the disease that ultimately kills a person; it is the inability to fight off something as minor as the common cold. • www.dellpassovoy.com/Viruses.ppt - Mirip To date, there has been no discovery of a treatment which is capable of either totally prohibiting or eliminating the HIV/AIDS virus and is effective and useful for humans

27. These are different targets where one could Fight and possibly kill the HIV virus. 1. Binding of HIV to membrane of target cells. 2. Inhibiting the reverse transcriptase (converts HIV RNA to DNA) 3. Inhibiting the enzyme that degrades HIV RNA after conversion to DNA 4. Inhibiting the actions of HIV integrase that integrates HIV DNA into host cells 5. Inhibiting the expression of HIV genetic material once integrated into the host cell 6. Inhibiting protease that splices proteins for assembling new virus particles.

28. There are three drug types existing that are used in treating HIV. • Nucleoside reverse transcriptase inhibitors (Example- AZT, etc). These are the most well developed and widely studied medications. These function by interfering with the reverse transcriptase enzyme while the viral RNA is being transcribed to DNA. The virus functions by causing a termination in the chain of nucleic acid as it is being assembled. Early treatment with AZT has been proved to establish longer survival times except long term use can result in horrible long term irreversible side effects as well as short term side effects such as headache, diaherra, muscle pain, fever, insomnia. • Non-nucleoside reverse transcriptase inhibitors. (Example- Delavidine, nevirapine) These function similar to the reverse transcriptase inhibitors except they terminate the nucleic acid chain at an earlier site.

29. Protease inhibitors, which are called AIDS cocktails. (Example-Saquinavir, Ritonavir, Crxivan) They disrupt the HIV replication cycle as well, except at a different point. Protease is another enzyme essential to the replication of the virus. Without it, the formation and organization of virus proteins remains incomplete, rendering the virus particles inactive. • Nucleocides interfere with the virus nucleotides before they enter the nucleus of the cell and are processed as proteins. Protease Inhibitors, on the other hand, interfere with the assembly of the proteins after they are processed. The problem with all of these drugs is that the virus changes so often and so rapidly. It is almost a guess as to how the virus will change, this is the reason why "drug cocktails" are used most often to fight the HIV virus.

30. The main problem with the current treatments is that the HIV virus mutates quickly and that can render the treatments ineffective. Rather than trying to conquer the constantly changing, our idea was to cover the unchanging with a protective barrier. We ideally would like to create a protein, with similar characteristics to a gp120 protein that would react with the CD4 cells in our body thus preventing the HIV virus from reacting with the CD4 and depositing nucleic acid. • The latest research on HIV treatment has to do with the subject of inhibiting binding to the human cells. The proteins found in starfish inhibited the infection of human T-lymphoblastoid cells by HIV-1 in vitro with an EC50 of approximately 4 ng/mL. However, it was toxic in the human body. The new challenge is to find something that will not be toxic in humans but contains the same proteins, thus performing the same duties as the starfish protein on human CD4 cells.

31. What is not known • It is not known how this starfish protein can be used in humans as it is because because it is toxic. It is also unknown how to make it non-toxic. Research should be done on how this protein could be altered, hopefully at just one point in the genetic code, to account for the difference. • While researching nutritional information on HIV it was found that those patients who had a healthier diet had an increased number of t-cells. Also a startling phenomenon was discovered--(suspense rises) there is an entire body protein turn-over--this means liberated amino acids were converted to synthesize new protein. Is it possible that the body is trying to produce new gp 120 proteins to block the CD4 receptor sites? • Also, another amazing trend has appeared... the t-cell count for HIV infected patients decreases rapidly for 3 months and increases back to normal levels in the following three months.

32. What are vaccines?www.scripps.edu/community/lessonplans/Gibson%201.ppt - Amerika Serikat • A vaccine is a substance that stimulates the body’s immune response. • The goal of vaccination is to prevent or control an infection. • A vaccine is an antigenic preparation used to establish immunity to a disease. • Vaccines can be prophylactic (e.g. to prevent or ameliorate the effects of a future infection by any natural or "wild" pathogen), or therapeutic (e.g. vaccines against cancer are also being investigated; see cancer vaccine

34. There are four types of traditional vaccines • Vaccines containing killed microorganisms - these are previously virulent micro-organisms that have been killed with chemicals or heat. Examples are vaccines against flu, cholera, , and hepatitis A. • Vaccines containing live, attenuated microorganisms - these are live micro-organisms that have been cultivated under conditions that disable their virulent properties. They typically provoke more durable immunological responses and are the preferred type for healthy adults. Examples include yellow fever, measles, rubella, and mumps. • Toxoids - these are inactivated toxic compounds from micro-organisms in cases where these (rather than the micro-organism itself) cause illness. Examples of toxoid-based vaccines include tetanus and diphtheria. • Subunit - rather than introducing an inactivated or attenuated micro-organism to an immune system, a fragment of it can create an immune response. Characteristic examples include the subunit vaccine against HBV that is composed of only the surface proteins of the virus (produced in yeast) and the virus-like particle (VLP) vaccine against human papillomavirus (HPV) that is composed of the viral major capsid protein.

35. An Innovative vaccines are also in development and in use: • Conjugate - certain bacteria have polysaccharide outer coats that are poorly immunogenic. By linking these outer coats to proteins (e.g. toxins), the immune system can be led to recognize the polysaccharide as if it were a protein antigen. This approach is used in the Haemophilus influenzae type B vaccine. • Recombinant Vector - by combining the physiology of one micro-organism and the DNA of the other, immunity can be created against diseases that have complex infection processes • DNA vaccination - in recent years a new type of vaccine, created from an infectious agent's DNA called DNA vaccination, has been developed. It works by insertion (and expression, triggering immune system recognition) into human or animal cells, of viral or bacterial DNA. Some cells of the immune system that recognize the proteins expressed will mount an attack against these proteins and cells expressing them. Because these cells live for a very long time, if the pathogen that normally expresses these proteins is encountered at a later time, they will be attacked instantly by the immune system. One advantage of DNA vaccines is that they are very easy to produce and store. As of 2006, DNA vaccination is still experimental, but shows some promising results.

36. Influenza Virus • Influenza A virus is essentially an avian virus that has "recently" crossed into mammals. Birds have the greatest number and range of influenza strains. Avian haemagglutinins sometimes appear in pig human and horse influenza strains. • Every now and then (10 - 15 years) a major new pandemic strain appears in man, with a totally new HA and sometimes a new NA as well (antigenic shift). This variant causes a major epidemic around the world (pandemic). • Over the subsequent years this strain undergoes minor changes (antigenic drift) every two to three years, probably driven by selective antibody pressure in the populations of humans infected. • The avian influenzavirus subtypes that have been confirmed in humans, ordered by the number of known human deaths, are: H1N1 caused Spanish flu, H2N2 caused Asian Flu, H3N2 caused Hong Kong Flu, H5N1 is the current pandemic threat, H7N7, H9N2, H7N2, H7N3.

37. The virion is generally rounded but may be long and filamentous.A single-stranded RNA genome is closely associated with a helical nucleoprotein (NP), and is present in eight separate segments of ribonucleoprotein (RNP), each of which has to be present for successful replication. The segmented genome is enclosed within an outer lipoprotein envelope. An antigenic protein called thematrix protein (MP 1)lines the inside of the envelope and and is chemically bound to the RNP. The envelope carries two types of protruding spikes. One is a box-shaped protein, called the neuraminidase (NA), of which there are nine major antigenic types, and which has enzymic properties as the name implies. • The other type of envelope spike is a trimeric protein called the haemagglutinin (HA) (illustrated on the right) of which there are 15 major antigenic types. The haemagglutinin functions during attachment of the virus particle to the cell membrane, and can combine with specific receptors on a variety of cells including red blood cells. The lipoprotein envelope makes the virion rather labile - susceptible to heat, drying, detergents and solvents.

38. Neuraminidase Haemaglutinin RNA M protein Only in Type A

40. Avian influenza virus spreads in the air and in manure and survives longer in cold weather. It can also be transmitted by contaminated feed, water, equipment and clothing; however, there is no evidence that the virus can survive in well cooked meat. The incubation period is 3 to 5 days. Symptoms in animals vary, but virulent strains can cause death within a few days. • H5N1 H5N1 is a highly pathogenic form of avian influenzavirus. Since 1997, outbreaks of H5N1 flu have caused the death or culling of tens of millions of birds. Over 100 people have been infected by H5N1, with a mortality rate of over 50%. H5N1 has been the focus of much concern amid warnings that the H5N1 strain will likely evolve into a form that causes a global human pandemic with a very high mortality rate. As of November 1, 2005, 122 cases of infections in humans, resulting in 62 deaths, have been confirmed outside of China. • Antiviral drugs such as oseltamivir, zanamivir and amantadine are sometimes effective in both preventing and treating the infection. Countries have been stockpiling olestamivir, but may shift towards zanamivir due to issue, which reported oseltamivir resistant strains of avian flu in Vietnam. Flu vaccines, however, take at least four months to produce and must be prepared for each subtype.