Lesson 12

1. Lesson 12 Virus Biology

2. Virus: definition • A virus (from the Latin virus meaning "toxin" or "poison"), is a sub-microscopic infectious agent that is unable to grow or reproduce outside a host cell. Each viral particle, or virion, consists of genetic material, DNA or RNA, within a protective protein coat called a capsid. The capsid shape varies from simple helical and icosahedral (polyhedral or near-spherical) forms, to more complex structures with tails or an envelope. Viruses infect cellular life forms and are grouped into animal, plant and bacterial types, according to the type of host infected. • Biologists debate whether or not viruses are living organisms. Some consider them non-living as they do not meet all the criteria used in the common definitions of life. For example, unlike most organisms, viruses do not have cells. However, viruses have genes and evolve by natural selection. Others have described them as organisms at the edge of life. Viral infections in human and animal hosts usually result in an immune response and disease. Often, a virus is completely eliminated by the immune system. 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.

3. Viruses: an Overview • Viruses are parasites that reproduce inside the cells of the organisms they infect. When a virus infects a cell, it reprograms that cell's normal biochemical processes thereby forcing the cell to make thousands of identical copies of the infecting virus by using the cell's protein, DNA and RNA building blocks. This process usually kills the infected cell. • Viruses are grouped depending on the type of organism they infect. Therefore viruses are either animal, plant or bacterial types. They are so small that it would take 30,000 to 750,000 of them, side by side, to cover 1 centimetre. The origins of viruses is unclear; some may have evolved from plasmids or transposons, which are stretches of DNA that can exist separately from chromosomes. Other viruses may have evolved from bacteria. • A virus consists of two parts: the genetic material, a nucleic acid that contains all the information necessary for the production of new virus particles, and a protein coat that protects this genetic material. The coat is called a capsid and can either be a simple container, or a much more complex structure, containing enzymes. Capsid shapes vary from simple helical and icosahedral (polyhedral or near-spherical) forms, to more complex structures with tails or an envelope. • Plant, animal and bacterial cells all contain two types of nucleic acid. The genetic information needed to make new proteins is always encoded in DNA: this code is translated into a specific type of RNA called messenger RNA that carries the information to the parts of the cell that make proteins. In contrast, each type of virus only contains one type of nucleic acid, either DNA or RNA. • Within cells there are structures called organelles, each organelle has one or more important functions. These organelles along with cell enzymes are needed for metabolism and life. Viruses do not have organelles and must use those within a cell to reproduce. Outside of a host cell, viruses are completely inactive and are considered neither living nor dead. Indeed, it is controversial whether viruses are actually living organisms. Some consider them non-living since they do not meet all the criteria of the definition of life. However, viruses do have genes and evolve over time. • Viral infections in human and animals usually cause an immune response and disease. Often, a virus is completely eliminated by the immune system. Antibiotics have no effect on viruses, but antiviral drugs have been developed to treat life-threatening infections. Vaccines that produce lifelong immunity can prevent virus infections. • Viruses spread in many different ways. Viruses of plants are often spread from plant to plant by insects acting as vectors. Specific virus infections of animals and humans are also spread by biting insects. But not all viruses can be spread in this way. Each species of virus relies on a specific method. Some, such as influenza are carried by the air and are spread by coughing and sneezing. Others, such as norovirus and rotavirus contaminate food or water. HIV is one of several viruses that are transmitted during sex.

4. Virus classification • The International Committee on Taxonomy of Viruses (ICTV) developed the current classification system and put in place guidelines that put a greater weighting on certain virus properties to maintain family uniformity. A universal system for classifying viruses, and a unified taxonomy, has been established since 1966. In determining order, taxonomists should consider the type of nucleic acid present, whether the nucleic acid is single- or double-stranded, and the presence or absence of an envelope. After these three main properties, other characteristics can be considered: the type of host, the capsid shape, immunological properties and the type of disease it causes. The system makes use of a series of ranked taxons. The general structure is as follows: • Order (-virales) • Family (-viridae) • Subfamily (-virinae) • Genus (-virus) • Species (-virus) • The recognition of orders is very recent; to date, only three have been named, and most families remain unplaced. The committee does not formally distinguish between subspecies, strains, and isolates. In total there are three orders, 56 families, nine subfamilies, and 233 genera. ICTV recognizes about 1,550 virus species, but about 30,000 virus strains and isolates are being tracked by virologists.

5. Baltimore Classification • The Nobel Prize-winning biologist David Baltimore devised the Baltimore classification system. The ICTV classification system is used in conjunction with the Baltimore classification system in modern virus classification. • The Baltimore classification of viruses is based on the mechanism of mRNA production. Viruses must generate positive strand mRNAs from their genomes to produce proteins and replicate themselves, but different mechanisms are used to achieve this in each virus family. This classification places viruses into seven groups. • I: dsDNA viruses • II: ssDNA viruses • III: dsRNA viruses • IV: (+)ssRNA viruses • V: (-)ssRNA viruses • VI: ssRNA-RT viruses • VII: dsDNA-RT viruses • ds: double strand • ss: single strand • RT: reverse transcribing Computer reconstruction of a rotavirus particle

6. Baltimore Classification • The Baltimore classification is a virus classification system which groups viruses into families depending on their type of genome (DNA, RNA, single-stranded (ss), double-stranded (ds) etc.) and their method of replication.

7. Structure of viruses • A complete virus particle, known as a virion, consists of nucleic acid surrounded by a protective coat of protein called a capsid. Viruses can have a lipid "envelope" derived from the host cell membrane. Most DNA viruses do not have an envelope but most RNA viruses are enveloped. • A capsid is made from proteins encoded by the viral genome and its shape serves as the basis for morphological and antigenic distinction. Virally coded protein subunits will self-assemble to form a capsid, generally requiring the presence of the virus genome. However, complex viruses code for proteins which assist in the construction of their capsid. Proteins associated with nucleic acid are known as nucleoproteins, and the association of viral capsid proteins with viral nucleic acid is called a nucleocapsid. • Viruses are some of the smallest infectious agents and the majority of them can only be seen by electron microscopy. Most viruses are around 100 nm in diameter but their sizes range from 20 to 300nm. They are so small that it would take 30,000 to 750,000 of them, side by side, to cover 1 cm.

8. Helical viruses • Helical capsids are composed of a single type of subunit stacked around a central axis to form a helicalstructure which may have a central cavity, or hollow tube. This arrangement results in rod-shaped or filamentous virions: these can be anything from short and highly rigid, to long and very flexible. The genetic material, generally single-stranded RNA, but ssDNA in some cases, is bound into the protein helix, by interactions between the negatively-charged nucleic acid and positive charges on the protein. Overall, the length of a helical capsid is related to the length of the nucleic acid contained within it and the diameter is dependent on the size and arrangement of protomers. The well-studied Tobacco mosaic virus (see Figure) is an example of a helical virus.

9. Icosahedron viruses • Icosahedral* capsid symmetry results in a spherical appearance of viruses at low magnification but actually consists of capsomers arranged in a regular geometrical pattern, similar to a soccer ball, hence they are not truly "spherical". Capsomers are ring shaped constructed from five to six copies of protomers. These associate via non-covalent bonding to enclose the viral nucleic acid, though generally less intimately than helical capsids, and may involve one or more protomers. • Icosahedral architecture is the most efficient way of creating an enclosed robust structure from multiple copies of a single protein. The number of proteins required to form a spherical virus capsid is denoted by the T-number, where 60×t proteins are necessary. In the case of the hepatitis B virus the T-number is 4, and 240 proteins assemble to form the capsid. • Figure: Adenovirus • * is any polyhedron having 20 faces, but usually a regular icosahedron is implied, which has equilateral triangles as faces.

10. Enveloped viruses • Viruses are able to envelope themselves in a modified form of one of the cell membranes either the outer membrane surrounding an infected host cell, or from internal membranes such as nuclear membrane or endoplasmic reticulum, thus gaining an outer lipid bilayer known as a viral envelope. This membrane is studded with proteins coded for by the viral genome and host genome; however the lipid membrane itself and any carbohydrates present are entirely host-coded. The Influenza virus and HIV use this strategy. • The viral envelope can give a virion a few distinct advantages over other capsid-only virions, such as protection from enzymes and certain chemicals. The proteins in it can include glycoproteins functioning as receptor molecules, allowing host cells to recognize and bind these virions, resulting in the possible uptake of the virion into the cell. Most enveloped viruses are dependent on the envelope for infectivity. • Fig: Herpes zoster virus

11. Complex viruses I • These viruses possess a capsid which is neither purely helical, nor purely icosahedral, and which may possess extra structures such as protein tails or a complex outer wall. Some bacteriophages (Figure) have a complex structure consisting of an icosahedral head bound to a helical tail, the latter of which may have a hexagonal base plate with protruding protein tail fibres.

12. Complex viruses II • The Poxviruses (Figure) are large, complex viruses which have an unusual morphology. The viral genome is associated with proteins within a central disk structure known as a nucleoid. The nucleoid is surrounded by a membrane and two lateral bodies of unknown function. The virus has an outer envelope with a thick layer of protein studded over its surface. The whole particle is slightly pleiomorphic, ranging from ovoid to brick shape.

13. Virus genes • There are two kinds of genetic material: deoxyribonucleic acid (DNA) and ribonucleic acid (RNA). The biological information contained in an organism is encoded in its DNA or RNA sequence. Most organisms use DNA, but many viruses (e.g., retroviruses) have RNA as their genetic material. The DNA and RNA of viruses consists of either a single strand or a double helix. • Viruses, although simple compared to cell-based organisms, are very efficient at reproducing. They have only a few genes compared to humans who have 20,000–25,000. For example, Influenza virus has only 8 genes and rotavirus has 11. These genes encode structural proteins that form the virus particle, or non-structural proteins, that are only found in cells infected by the virus. • Most viruses produce a protein that is an enzyme called a polymerase. Polymerase is used to make new copies of the viral DNA or RNA. Often this protein is also a structural protein that forms part of the virus particle. These polymerase enzymes are often much more efficient than their counterparts produced by the host cell. • In some species of virus the DNA or RNA is not a continuous molecule but is split into several separate strands. These are called segmented genomes. The Influenza virus genome is made up from 8 separate segments of RNA. When two different strains of influenza virus infect the same cell, these segments can mix and produce new strains of the virus, this is called reassortment.

14. Genomic diversity among viruses • An enormous variety of genomic structures can be seen among viral species; as a group they contain more structural genomic diversity than the entire kingdoms of either plants, animals, or bacteria. • A virus may employ either DNA or RNA as the nucleic acid. Rarely do they contain both, however cytomegalovirus is an exception to this, possessing a DNA core with several mRNA segments. By far most viruses have RNA. Plant viruses tend to have single-stranded RNA and bacteriophages tend to have double-stranded DNA. Some virus species possess abnormal nucleotides, such as hydroxymethylcytosine instead of cytosine, as a normal part of their genome.

15. Virus Genome I • Shape • Viral genomes may be circular, such as polyomaviruses, or linear, such as adenoviruses. The type of nucleic acid is irrelevant to the shape of the genome. Among RNA viruses, the genome is often divided up into separate parts within the virion and are called segmented. Double-stranded RNA genomes and some single-stranded RNA genomes are segmented. Each segment often codes for one protein and they are usually found together in one capsid. Every segment is not required to be in the same virion for the overall virus to be infectious, as demonstrated by the brome mosaic virus. • Strandedness • A viral genome, irrespective of nucleic acid type, may be either single-stranded or double-stranded. Single-stranded genomes consist of an unpaired nucleic acid, analogous to one-half of a ladder split down the middle. Double-stranded genomes consist of two complementary paired nucleic acids, analogous to a ladder. Viruses, such as those belonging to the Hepadnaviridae, contain a genome which is partially double-stranded and partially single-stranded. Viruses that infect humans include double-stranded RNA (e.g. Rotavirus), single-stranded RNA (e.g. Influenza virus), single-stranded DNA (e.g. Parvovirus B19) and double-stranded DNA (Herpes virus). • Sense • For viruses with RNA as their nucleic acid, the strands are said to be either positive-sense (called the plus-strand) or negative-sense (called the minus-strand), depending on whether it is complementary to viral mRNA. Positive-sense viral RNA is identical to viral mRNA and thus can be immediately translated by the host cell. Negative-sense viral RNA is complementary to mRNA and thus must be converted to positive-sense RNA by an RNA polymerase before translation. DNA nomenclature is similar to RNA nomenclature, in that the coding strand for the viral mRNA is complementary to it (-), and the non-coding strand is a copy of it (+).

16. Virus Genome II • Genome size • Genome size in terms of the weight of nucleotides varies between species. The smallest genomes code for only four proteins and weigh about 106 Daltons, the largest weigh about 108 kDaltons and code for over one hundred proteins. RNA viruses generally have smaller genome sizes than DNA viruses due to a higher error-rate when replicating, resulting in a maximum upper size limit. Beyond this limit, errors in the genome when replicating render the virus useless or uncompetitive. To compensate for this, RNA viruses often have segmented genomes where the genome is split into smaller molecules, thus reducing the chance of error. In contrast, DNA viruses generally have larger genomes due to the high fidelity of their replication enzymes. • Gene reassortment • There is an evolutionary advantage in having a segmented genome. Different strains of a virus with a segmented genome, from a pig or a bird or a human for example, such as Influenza virus, can shuffle and combine with other genes producing progeny viruses or (offspring) that have unique characteristics. This is called reassortment or viral sex. This is one reason why Influenza virus constantly changes. • Genetic recombination • Genetic recombination is the process by which a strand of DNA is broken and then joined to the end of a different DNA molecule. This can occur when viruses infect cells simultaneously and studies of viral evolution have shown that recombination has been rampant in the species studied. Recombination is common to both RNA and DNA viruses. • Genetic change • Viruses undergo genetic change by several mechanisms. These include a process called genetic drift where individual bases in the DNA or RNA mutate to other bases. Most of these point mutations are silent in that they do not change the protein that the gene encodes, but others can confer evolutionary advantages such as resistance to antiviral drugs. Antigenic shift is where there is a major change in the genome of the virus. This occurs as a result of recombination or reassortment (see above). When this happens with influenza viruses, pandemics may result. By genome rearrangement the structure of the gene changes although no mutations have necessarily occurred. • RNA viruses are much more likely to mutate than DNA viruses for the reasons outlined above. Viruses often exist as quasispecies or swarms of viruses of the same species but with slightly different genome nucleoside sequences. Such quasispecies are a prime target for natural selection.

17. Virus envelope structure • Many viruses (e.g. influenza and many animal viruses) have viral envelopes covering their protein capsids. The envelopes are typically derived from portions of the host cell membranes (phospholipids and proteins), but include some viral glycoproteins. Functionally, viral envelopes are used to help viruses enter host cells. Glycoproteins on the surface of the envelope serve to identify and bind to receptor sites on the host's membrane. The viral envelope then fuses with the host's membrane, allowing the capsid and viral genome to enter and infect the host. Cytomegalievirus (CMV); Herpes

18. Replication of viruses • Viral populations do not grow through cell division, because they are acellular; instead, they use the machinery and metabolism of a host cell to produce multiple copies of themselves. A virus can still cause degenerative effects within a cell without causing its death; collectively these are termed cytopathic effects.

19. Viral life cycle • The life cycle of viruses differs greatly between species (see below) but there are basic stages in the life cycle of viruses: • A virus attaches to the host cell and enters endocytosis. • The capsid protein dissociates and the viral RNA is transported to the nucleus. • In the nucleus, the viral polymerase complexes transcribe and replicate the RNA. • Viral mRNAs migrate to cytoplasm where they are translated into protein. • Then the newly synthesized virions bud from infected cell.

20. Viral life cycle • * Attachment is a specific binding between viral capsid proteins and specific receptors on the host cellular surface. This specificity determines the host range of a virus. For example, the human immunodeficiency virus (HIV) infects only human T cells, because its surface protein, gp120, can interact with CD4 and receptors on the T cell's surface. This mechanism has evolved to favour those viruses that only infect cells that they are capable of replicating in. Attachment to the receptor can induce the viral-envelope protein to undergo changes that results in the fusion of viral and cellular membranes. • * Penetration: following attachment, viruses enter the host cell through receptor mediated endocytosis or membrane fusion. • * Uncoating is a process in which 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 (see above), viral protein synthesis and assembly of viral proteins and viral genome replication. • * Following the assembly of the virus particles post-translational modification of the viral proteins often occurs. In viruses such as HIV, this modification, (sometimes called maturation), occurs after the virus has been released from the host cell. • * Viruses are released from the host cell by lysis . Enveloped viruses (e.g., HIV) typically are released from the host cell by budding. During this process, the virus acquires its phospholipid envelope which contains embedded viral glycoproteins.

21. Virus protein synthesis • Cells make new protein molecules from amino acid building blocks based on information encoded in RNA. Protein synthesis generally consists of two major steps: transcription and translation. Transcription is the process where genetic information in DNA is used to produce messenger RNA which migrates through the cell. The mRNA molecules bind to protein-RNA complexes called ribosomes where they are used to make proteins. This is called translation. The protein amino acid sequence is based on the mRNA base sequence. • Virus particles are made from several proteins that they cannot produce by themselves, therefore they rely on the host cell to make them. Viruses do this in many different ways, however, eventually all of them produce mRNA. In some RNA viruses the viral genome RNA functions directly as mRNA without further modification. For this reason, these viruses are called positive-sense RNA viruses. • In other RNA viruses, the genomic RNA is a complementary copy of mRNA and these viruses rely on the cell's or their own enzyme to make mRNA. These are called negative-sense RNA viruses. In viruses made of DNA the method of mRNA production is similar to a cell. Retroviruses are very different, they are based on RNA, but inside the host cell a DNA copy of their RNA is made. This DNA is then incorporated into the host's and copied into mRNA by the cell's normal pathways.

22. Viral proteins: influenza virus • The influenza A virus particle or virion is 80–120 nm in diameter and usually roughly spherical, although filamentous forms can occur. Unusually for a virus, the influenza A genome is not a single piece of nucleic acid; instead, it contains eight pieces of segmented negative-sense RNA (13.5 kilobases total), which encode 11 proteins (HA (hemagglutinin), NA (neuraminidase), NP (nucleoprotein), M1, M2, NS1, NS2(NEP), PA, PB1, PB1-F2, PB2). • The best-characterised of these viral proteins are hemagglutinin and neuraminidase, two large glycoproteins found on the outside of the viral particles. Neuraminidase is an enzyme involved in the release of progeny virus from infected cells, by cleaving sugars that bind the mature viral particles. By contrast, hemagglutinin is a lectin that mediates binding of the virus to target cells and entry of the viral genome into the target cell. The hemagglutinin (HA or H) and neuraminidase (NA or N) proteins are targets for antiviral drugs. These proteins are also recognised by antibodies, i.e. they are antigens. The responses of antibodies to these proteins are used to classify the different serotypes of influenza A viruses, hence the H and N in H5N1.

23. Viruses and disease • Human diseases caused by viruses include the common cold, the flu, chickenpox and cold sores. Serious diseases such as Ebola, AIDS and influenza are caused by viruses. The ability of viruses to cause disease is called virulence and the mechanism is called pathogenesis. Viruses cause different diseases depending on the types of cell that they infect. Some viruses can cause life-long or chronic infections where the viruses continue to replicate in the body despite the hosts' defense mechanisms. This is common in Hepatitis B virus and Hepatitis C Virus infections. People chronically infected with Hepatitis B virus are known as carriers who serve as reservoirs of infectious virus. In some populations, if there is a high proportion of carriers, a disease is said to be endemic. • There are many ways in which viruses spread from host to host but each species of virus uses only one or two. Many viruses that infectplants are carried by insects called vectors. Common vectors include Bacteria, Fungi, Nematodes, arthropods and arachnids. Some viruses that infect animals and humans are also spread by biting insects, but direct animal-to-animal, person-to-person or animal-to-person transmission is more common. Some virus infections are spread by contaminated food and water, (Norovirus and Rotavirus), others are airborne, (Influenza virus). Viruses such as HIV, Hepatitis B and Hepatitis C are often transmitted by unprotected sex or contaminated hypodermic needles. It is important to know how each different kind of virus is spread in order to prevent infections and epidemics.

24. Comparison table of clinically important virus families and species

25. Virus vaccines • Vaccination is a way of preventing infections by viruses. Vaccines were used to prevent viral infections long before the discovery of the actual viruses. Their use has resulted in a dramatic decline in morbidity (illness) and mortality (death) associated with viral infections such as polio, measles, mumps and rubella. Smallpox infections have been eradicated. Currently vaccines are available to prevent over thirteen viral infections of humans and more are used to prevent viral infections of animals. • Vaccines may consist of live or killed viruses. Live vaccines contain weakened forms of the virus that are no longer virulent. These are referred to as attenuated vaccines. Live vaccines can be dangerous when given to immuno-compromised people i.e. people with weak immunity, because in these people, the weakened virus can cause the original disease. Biotechnology and genetic engineering techniques are used to produce subunit vaccines. These vaccines use only the capsid proteins of the virus. Hepatitis B vaccine is an example of this type of vaccine. Subunit vaccines are safe for immunocompromised patients because they cannot cause the disease.

26. Antiviral drugs • Over the past twenty years the development of antiviral drugs has increased rapidly. This has been driven by the AIDS epidemic. Antiviral drugs are often nucleoside analogues, (fake DNA building blocks), which are incorporated into the viral DNA during its replication. DNA, and thus virus replication is then halted because these analogues lack the hydroxyl groups which along with phosphorus atoms, link together to form the strong "backbone" of the DNA molecule. This is called DNA chain termination. Examples of nucleoside analogues are aciclovir for Herpes virus infections and lamivudine for HIV and Hepatitis B virus infections. Aciclovir, is one of the oldest and most frequently prescribed antiviral drugs. Other antiviral drugs target different stages of the viral life cycle. HIV is dependent on a proteolytic enzyme called the HIV-1 protease for it to become fully infectious. There is a class of drugs called protease inhibitors which have been designed to inactivate the enzyme. • Hepatitis C is caused by an RNA virus. In 80% of people infected, the disease is chronic and without treatment they remain infectious for the rest of their lives. However, there is now an effective treatment using the nucleoside analogue drug ribavirin combined with interferon. The treatment of chronic carriers of the Hepatitis B virus by using a similar strategy using lamivudine is being developed.

27. Viruses in life sciences and medicine • Viruses are important to the study of molecular and cellular biology as they provide simple systems that can be used to manipulate and investigate the functions of cells. The study and use of viruses have provided valuable information about aspects of cell biology. For example, viruses have been useful in the study of genetics and helped our understanding of the basic mechanisms of molecular genetics, such as DNA replication, trans-cription, RNA processing, translation, protein transport, and immunology. • Geneticists often use viruses as vectors to introduce genes into cells that they are studying. This is useful for making the cell produce a foreign substance, or to study the effect of introducing a new gene into the genome. In similar fashion, virotherapy uses viruses as vectors to treat various diseases, as they can specifically target cells and DNA. It shows promising use in the treatment of cancer and in gene therapy. Eastern European scientists have used phage therapy as an alternative to antibiotics for some time, and interest in this approach is increasing, due to the high level of antibiotic resistance now found in some pathogenic bacteria. • Granulosis (GV) and nucleo-polyhedrosis viruses (NPV) may also be used as biological insecticides (e.g. Cydia pomonella granulovirus).

28. Viruses in materials science and nanotechnology • Current trends in nanotechnology promise to make much more versatile use of viruses. From the viewpoint of a materials scientist, viruses can be regarded as organic nanoparticles. Their surface carries specific tools designed to cross the barriers of their host cells. The size and shape of viruses, and the number and nature of the functional groups on their surface, is precisely defined. As such, viruses are commonly used in materials science as scaffolds for covalently linked surface modifications. A particular quality of viruses is that they can be tailored by directed evolution. The powerful techniques developed by life sciences are becoming the basis of engineering approaches towards nanomaterials, opening a wide range of applications far beyond biology and medicine. • Because of their size, shape, and well-defined chemical structures, viruses have been used as templates for organizing materials on the nanoscale. Recent examples using Cowpea Mosaic Virus (CPMV) particles to amplify signals in microarray based sensors. In this application, the virus particles separate the fluorescent dyes used for signaling in order to prevent the formation of non-fluorescent dimers that act as quenchers. Another example is the use of CPMV as a nanoscale breadboard for molecular electronics. In April 2006, scientists at the Massachusetts Institute of Technology (MIT) created nanoscale metallic wires using a genetically-modified virus. The MIT team was able to use the virus to create a working battery with an energy density up to three times more than current materials. The potential exists for this technology to be used in liquid crystals, solar cells, fuel cells, and other electronics in the future.

29. Virus Weapons • The ability of viruses to cause devastating epidemics in human societies has led to the concern that viruses could be weaponised for biological warfare. Further concern was raised by the successful recreation of the infamous 1918 influenza virus in a laboratory. The smallpox virus devastated numerous societies throughout history before its eradication. It currently exists in several secure laboratories in the world, and fears that it may be used as a weapon are not totally unfounded. The vaccine for smallpox is not safe, and during the years before the eradication of smallpox disease more people became seriously ill as a result of vaccination than did people from smallpox and smallpox vaccination is no longer universally practiced. Thus, the modern global human population has almost no established resistance to smallpox; if it were to be released, a massive loss of life could be sustained before the virus is brought under control.