6 Viruses and Other Acellular Infectious Agents

Prescott’s Microbiology, 9th Edition

CHAPTER OVERVIEW

Viruses are small, acellular entities that usually possess only a single type of nucleic acid and must use the metabolic machinery of a living host in order to reproduce. Viruses have been and continue to be of tremendous importance because many human diseases have a viral etiology. The study of viruses has contributed greatly to our knowledge of molecular biology, and the blossoming field of genetic engineering is based on discoveries in the field of virology. This chapter focuses on the general properties of viruses, their structure, reproduction, infectivity, and cultivation. The discussion concludes with the features of viroids, virusoids, and prions.

LEARNING OUTCOMES

After reading this chapter you should be able to:

• define the terms virology, bacteriophages and phages • list organisms that are hosts to viruses • state the size range of virions • identify the parts of a virion and describe their function • distinguish enveloped from noneveloped viruses • describe the types of capsid symmetry • describe the five steps common to the life cycles of all viruses • discuss the roles of receptors, capsid proteins, and envelope proteins in the life cycles of virues • describe the two most common methods for virion release from a host cell • compare and contrast the major steps of the life cycles of virulent phages and temperate phages • list examples of lysogenic conversion • differentiate among the types of viral infections of the eukaryotic cells • summarize the current understanding of how oncoviruses cause cancer • list the types of approaches used to cultivate viruses, noting which viruses are cultivated by each method • describe three direct counting methods and two indirect counting methods used to enumerate viruses • outline the events that lead to the formation of a plaque in a lawn of bacterial cells • distinguish lethal dose from infectious dose • describe the structure of a viroid and the discuss the practical importance of viroids • distinguish satellite viruses from satellite nucleic acids • describe prion structure and how prions are thought to replicate • list characteristics common to all animal diseases caused by prions • name at least two human diseases caused by prions • describe the mechanisms by which a prion protein might first appear in a human brain cell

CHAPTER OUTLINE

1 © 2014 by McGraw-Hill Education. This is proprietary material solely for authorized instructor use. Not authorized for sale or distribution in any manner. This document may not be copied, scanned, duplicated, forwarded, distributed, or posted on a website, in whole or part. Prescott’s Microbiology, 9th Edition

I. Viruses A. Viruses are acellular infectious agents that cannot reproduce outside of living cells; virology is the study of viruses B. Viruses can infect all types of cells but are specific for certain organisms; bacteriophages (phages) infect bacteria II. Virion Structure A. Virion size—ranges from 10 nm to 400 nm B. General structural properties 1. Nucleocapsid—the nucleic acid plus the surrounding capsid (protein coat that surrounds the genome); for some viruses this may be the whole virion; other viruses may possess additional structures 2. Three types of capsid symmetry: icosahedral, helical, complex 3. Viral capsids are constructed from many copies of one or a few types of proteins (protomers), which are assembled together with the viral genome 4. Enveloped viruses have an outer membranous layer (envelope) surrounding the nucleocapsid; naked (nonenveloped) viruses lack an envelope C. Helical capsids—hollow tube with a protein wall shaped as a helix or spiral; may be either rigid or flexible D. Icosahedral capsids—regular polyhedron with 20 equilateral triangular faces and 12 vertices; appears spherical; constructed of capsomers (ring or knob-shaped units), each usually made of five (pentamers or pentons) or six protomers (hexamers or hexons); viral capsids self-assemble E. Viruses with capsids of complex symmetry 1. Poxviruses are large (200 to 400 nm) with an ovoid exterior shape 2. Some bacteriophages have complex, elaborate shapes composed of heads (icosahedral symmetry) coupled to tails (helical symmetry); the structure of the tail regions is particularly variable; such viruses are said to have binal symmetry F. Viral envelopes and enzymes 1. Envelopes are membrane structures surrounding some (but not all) viruses a. Lipids and carbohydrates are usually derived from the host membranes b. Proteins are virus-specific c. Many have protruding glycoprotein spikes (peplomers) such as the enzymes neuraminidase and hemagglutinin 2. Although viruses lack true metabolism, they may carry one or more enzymes, many of which are involved in viral nucleic acid replication; in some cases the enzyme is capsid-associated, but in most cases the enzyme is located within the capsid G. Viral genomes 1. Viral genome may be either RNA or DNA, single- or double-stranded 2. DNA viruses a. Most use double-stranded DNA (dsDNA) as the genome; the DNA can be linear or circular b. Many have one or more unusual bases (e.g., hydroxymethylcytosine instead of cytosine) 3. RNA viruses—most have single-stranded RNA (ssRNA) as their genome; many have segmented genomes (genomes consisting of more than one molecule); each being unique and frequently encoding a single protein III. Viral Multiplication A. Common steps in viral life cycles 1. Attachment (adsorption) to the host cell 2. Entry of the nucelocapsid or viral genome into the host cell 3. Expression of viral genes in host cell (synthesis stage); virus takes control of cell's machinery and produces viral proteins and genomes 4. New virions self-assemble into mature virions 5. Virions released by budding or cell lysis B. Attachment (adsorption) 1. Virions attach to specific receptors, usually cell surface proteins or glycoproteins that are required by the cell for normal cell functioning 2. receptor distribution often varies over the surface of the host cells and lipid rafts seem to be involved in both virion entry and assembly 2 © 2014 by McGraw-Hill Education. This is proprietary material solely for authorized instructor use. Not authorized for sale or distribution in any manner. This document may not be copied, scanned, duplicated, forwarded, distributed, or posted on a website, in whole or part. Prescott’s Microbiology, 9th Edition

C. Entry into the host 1. Shortly after adsorption, viruses penetrate and enter the host cell; virus uncoating (removal of the capsid and release of viral nucleic acid) occurs for some virions during or shortly after penetration 2. There appear to be three common modes of entry a. Fusion of the viral envelope with the host cytoplasmic membrane results in deposition of the nucleocapsid core within the cell b. Endocytosis; lysosomal enzymes and low endosomal pH often trigger the uncoating process c. Some nonenveloped viruses directly inject their genome into the cytoplasm 3. Once in the cytoplasm the nucleic acid may function while still attached to capsid components or may only function after completion of uncoating D. Synthesis stage 1. Viral life cycles are specific for the type of viral genome; dsDNA viruses follow the normal flow of genetic information; RNA viruses need to carry enzymes for genome replication not found in the host 2. Expression of early viral genes is devoted to taking over the host cell; proteins synthesized late in the infection are usually involved in capsid synthesis and assembly E. Assembly 1. Capsid proteins are synthesized by host cell ribosomes under the direction of viral late genes, often as several subassemblies 2. Empty procapsids are produced and the nucleic acid is inserted within a "packasome" protein complex 3. Some viruses assemble in the cytoplasm, while others assemble in the nucleus F. Virion release 1. Nonenveloped viruses are often released when the host cell lyses 2. Enveloped viruses are usually released by the following mechanism a. Virus-encoded proteins are incorporated into the host cell’s plasma membrane (some viruses use the host’s nuclear membrane, endoplasmic reticulum, Golgi apparatus, or other membranes) b. The nucleocapsid buds outward, forming the envelope during release; actin cytoskeleton microfilaments can aid virion release IV.Types of Viral Infections A. Infections of bacterial and archaeal cells: lysis and lysogeny 1. Virulent phages are only capable of the lytic cycle, where multiplication begins once entering the host and continues until the host lyses 2. Temperate phages are capable of lysogeny, a nonlytic relationship with their hosts (virulent phages lyse their hosts), in addition to the lytic cycle a. In lysogeny, the viral genome (called a prophage) remains in the host (usually integrated into the host chromosome) but does not kill (lyse) the host cell; the cells are said to be lysogenic (or are called lysogens) b. It may switch to the lytic cycle at some later time; this process is called induction 3. Lysogenic conversion is a change that is induced in the host phenotype by the presence of a prophage, and that is not directly related to the completion of the viral life cycle; examples include: a. Modification of lipopolysaccharide structure in infected Salmonella b. Production of diphtheria toxin only by lysogenized strains of Corynebacterium diphtheriae 4. Most bacteriophages are temperate; it is thought that being able to lysogenize bacteria is advantageous; supporting this is the observation that certain conditions favor the establishment of lysogeny B. Infection of eukaryotic cells 1. Cytocidal infections lead to cell death; some viruses persist in the host for many years; in some cases degenerative changes occur in cells (cytopathic effects) without causing lysis C. Viruses and cancer

1. Cancer is a disease where there is abnormal cell growth (neoplasia) and spread of the abnormal cells throughout the body (metastasis); tumors are growths or lumps of tissue that can be benign (nonspreading) or malignant (cancerous) 2. Carcinogenesis is a group of complex, multistep processes that involve a triggering event and the activity of oncogenes; cells revert to less differentiated states (anaplasia) 3. Viruses have been implicated in a number of human cancers including, Epstein-Barr virus (Burkitt's lymphoma), human herpesvirus 8 (Kaposi's sarcoma), Hepatitis B and C (liver cancer), human papillomavirus (cervical cancer), human T-cell lymphotropic virus I (T-cell leukemia) 4. Viruses may cause cancer by a variety of mechanisms a. Viruses may carry one or more cancer-causing genes (oncogenes) b. Viruses may produce a regulatory protein that activates cell division c. Viruses may insert a promoter or enhancer that causes abnormal expression of a cellular proto-oncogene (a cellular gene that regulates cell growth and reproduction), and thereby deregulating cell growth d. Viral proteins can inhibit the activity of tumor suppressor proteins (e.g., Rb and p53) that regulate cell cycling and monitor or repair DNA damage V.Cultivation and Enumeration of Viruses A. Cultivation requires a suitable host since viruses cannot reproduce outside of cells B. Bacterial and archaeal viruses are usually cultivated in broth or agar cultures of suitable host cells; broth cultures are usually clear, while areas of localized destruction (plaques) form in bacterial lawns on agar plates C. Embryonated eggs have long been used to culture animal viruses D. In tissue (cell) culture monolayers of animal cells host viruses, resulting in plaques and cytopathic effects E. Plant viruses can be cultivated in plant tissue cultures, cultures of separated plant cells, plant protoplast cultures, or whole plants; in whole plants they may cause localized necrotic lesions or generalized symptoms of infection F. Virus assays 1. Particle counts a. Direct counts can be made with an electron or epifluorescence microscope b. Indirect counts can be made using methods such as hemagglutination (virus particles can cause red blood cells to clump together or agglutinate) 2. Measures of infectivity a. Plaque assays involve plating dilutions of virus particles on a lawn of host cells; clear zones result from viral damage to the cells; results are expressed as plaque-forming units (PFUs); similar measurements can be made by counting the number of pocks or necrotic lesions b. Infectious dose assays are an end-point method for determining the smallest amount of virus needed to cause a measurable effect, usually on 50% of the exposed target units;

results are expressed as infectious dose (ID50) or lethal dose (LD50) VI.Viroids and Satellites A. Viroids are circular ssRNA molecules that lack a capsid and cause a variety of plant diseases; the ssRNA molecules do not act as mRNAs (do not encode genes) and appear to be replicated by host RNA polymerases B. Pathogenicity of viroids may result from RNA silencing, where viroid RNA forms dsRNA by pairing with host mRNAs; these dsRNA are destroyed by host cell defenses and, hence, certain host genes are not translated, leading to disease C. Virusoids are circular ssRNA molecules (satellite RNAs) that encode proteins and need helper viruses for infectivity; the hepatitis D virusoid requires the help of the hepatitis B virus VII. Prions A. Prions are proteinaceous infectious particles (PrP) that are not associated with a nucleic acid genome B. Genes that encode PrP have been identified in normal animal tissue 1. It is hypothesized that abnormal PrP causes prion diseases by inducing a change from the normal conformation of the cellular PrP to the abnormal form

2. This new abnormal PrP then causes other normal cellular PrP molecules to change to the abnormal form C. Prions cause progressive, degenerative central nervous system disorders, including scrapie (sheep and goats), bovine spongiform encephalopathy (mad cow disease), Kuru (related to New Guinea tribal ritual cannibalism), Creutzfeldt-Jakob disease (CJD), fatal familial insomnia and Gerstmann- Strassler-Scheinker syndrome

CRITICAL THINKING

1. Viruses are thought by some scientists to be among the most primitive of living organisms. Others suggest that they are not living organisms. Take a stand on this debate and defend your position. What would be the most likely arguments from those taking the opposing viewpoint? How would you counter their arguments?

2. Consider the sequence shown below. It represents a segment of the mRNA produced by a virus. Suppose the virus was a double-stranded DNA virus; what would the corresponding genomic sequence be? Suppose the virus was a plus-strand RNA virus; what would the corresponding genomic sequence be? Finally, suppose the virus was a negative-strand RNA virus; what would the corresponding genomic sequence be?

5'CUGGAUCA3'

3. Describe some of the characteristics seen in bacteriophages compared to eukaryotic cell viruses. What do you think the difference in cellular structure between prokaryotes and eukaryotes plays in terms of these differences?

CONCEPT MAPPING CHALLENGE 5 © 2014 by McGraw-Hill Education. This is proprietary material solely for authorized instructor use. Not authorized for sale or distribution in any manner. This document may not be copied, scanned, duplicated, forwarded, distributed, or posted on a website, in whole or part. Prescott’s Microbiology, 9th Edition

Construct a concept map that describes the 5 stages of viral life cycles and illustrates some specific examples of how that stage is accomplished. Use the stages of the life cycle, any other concepts or terms you need, and your own linking words between each pair of concepts in your map.

6 © 2014 by McGraw-Hill Education. This is proprietary material solely for authorized instructor use. Not authorized for sale or distribution in any manner. This document may not be copied, scanned, duplicated, forwarded, distributed, or posted on a website, in whole or part.