Brief Contents

Preface to Second Edition xix Preface to First Edition xxi Abbreviations Used in This Book xxiii Greek Letters Used in This Book xxvii Color Coding for Molecules xxix

Chapter 1 and Their Importance 1 Chapter 2 Methods Used in 9 Chapter 3 Structure 27 Chapter 4 Virus Transmission 45 Chapter 5 Attachment and Entry of Viruses into Cells 55 Chapter 6 , , and Transport 65 Chapter 7 Virus Genome Replication 83 Chapter 8 Assembly and Exit of Virions from Cells 93 Chapter 9 Outcomes of Infection for the 101 Chapter 10 Classifi cation and Nomenclature of Viruses 115 Chapter 11 Herpesviruses (and Other dsDNA Viruses) 121 Chapter 12 Parvoviruses (and Other ssDNA Viruses) 135 Chapter 13 Reoviruses (and Other dsRNA Viruses) 145 Chapter 14 Picornaviruses (and Other Plus-Strand RNA Viruses) 155 Chapter 15 RhabdovirusesCOPYRIGHTED (and Other Minus-Strand RNA Viruses) MATERIAL 169 Chapter 16 Infl uenza Virus 183 Chapter 17 195 Chapter 18 Human Immunodefi ciency Viruses 207 Chapter 19 Hepadnaviruses (and Other Reverse-Transcribing DNA Viruses) 223 Chapter 20 Bacterial Viruses 237 Chapter 21 Origins and of Viruses 263 Chapter 22 Emerging Viruses 277

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Chapter 23 Viruses and Cancer 289 Chapter 24 Survival of Infectivity 301 Chapter 25 Virus 307 Chapter 26 Anti-viral Drugs 315 Chapter 27 327

Virologists’ Vocabulary 335 Index 347

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Preface to Second Edition xix Preface to First Edition xxi Abbreviations Used in This Book xxiii Greek Letters Used in This Book xxvii Color Coding for Molecules xxix

Chapter 1 Viruses and Their Importance 1 1.1 Viruses Are Ubiquitous on Earth 2 1.2 Reasons for Studying Viruses 3 1.2.1 Some viruses cause disease 3 1.2.2 Some viruses are useful 3 1.2.3 Virus studies have contributed to knowledge 4 1.3 The Nature of Viruses 4 1.3.1 Viruses are small particles 4 1.3.2 Viruses have 5 1.3.3 Viruses are parasites 6 1.3.4 Some viruses are dependent on other viruses 6 1.3.5 Are viruses living or nonliving? 6 1.4 The Remainder of the Book 7

Chapter 2 Methods Used in Virology 9 2.1 Introduction to Methods Used in Virology 11 2.2 Cultivation of Viruses 11 2.2.1 Animal culture 11 2.3 Isolation of Viruses 12 2.4 Centrifugation 14 2.4.1 Differential centrifugation 14 2.4.2 Density gradient centrifugation 14 2.5 Structural Investigations of Cells and Virions 15 2.5.1 Light microscopy 15 2.5.2 Electron microscopy 15 2.5.3 X-ray crystallography 16 2.6 Electrophoretic Techniques 16 2.7 Detection of Viruses and Virus Components 16 2.7.1 Detection of virions 16 2.7.2 Detection of infectivity using cell cultures 16 2.7.3 Detection of virus 18 2.7.4 Detection of virus nucleic acids 18

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2.8 Infectivity Assays 20 2.8.1 Quantitative assays 20 2.8.2 Quantal assays 21 2.8.3 One-step growth curve 22 2.9 Virus Genetics 23 2.9.1 Genome sequencing 23 2.9.2 Genome manipulation 24 2.9.3 Investigation of function and expression 24 2.10 Investigation of –Protein Interactions 25

Chapter 3 Virus Structure 27 3.1 Introduction to Virus Structure 28 3.2 Virus Genomes 28 3.2.1 Genome size 28 3.2.2 Secondary and tertiary structure 28 3.2.3 Modifi cations at the ends of virus genomes 29 3.2.4 non-covalently associated with virus genomes 32 3.2.5 Segmented genomes 33 3.2.6 Repeat sequences 33 3.3 Virus Proteins 33 3.4 35 3.4.1 Capsids with helical symmetry 35 3.4.2 Capsids with icosahedral symmetry 36 3.4.3 Conical and rod-shaped capsids 40 3.5 Virion Membranes 40 3.5.1 Enveloped virions 41 3.5.2 Virions with internal membranes 42 3.5.3 Membrane 42 3.6 Occlusion Bodies 42 3.7 Other Virion Components 43 3.7.1 Virus RNA in DNA viruses 43 3.7.2 Cell molecules in virions 43

Chapter 4 Virus Transmission 45 4.1 Introduction to Virus Transmission 46 4.1.1 Transmission via vectors: general principles 46 4.2 Transmission of Plant Viruses 47 4.3 Transmission of Vertebrate Viruses 49 4.3.1 Non- transmission of vertebrate viruses 49 4.3.2 Vector transmission of vertebrate viruses 50 4.3.3 Virus survival in a new vertebrate host 51 4.4 Transmission of Invertebrate Viruses 51 4.5 Permissive Cells 52

Chapter 5 Attachment and Entry of Viruses into Cells 55 5.1 Overview of Virus Replication 56 5.2 Animal Viruses 57

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5.2.1 Cell receptors and co-receptors 57 5.2.2 Virus attachment sites 58 5.2.3 Attachment of virions to receptors 58 5.2.4 Entry of animal viruses into cells 58 5.2.5 Intracellular transport 60 5.2.6 Genome uncoating 63 5.3 63

Chapter 6 Transcription, Translation, and Transport 65 6.1 Introduction to Transcription, Translation, and Transport 66 6.2 Transcription of Virus Genomes 66 6.2.1 Modifi cations to the central dogma 68 6.3 Transcription in 68 6.3.1 Promoters and enhancers 68 6.3.2 Transcription factors 70 6.3.3 Transcriptases 70 6.3.4 Capping transcripts 71 6.3.5 Polyadenylation of transcripts 72 6.3.6 Splicing transcripts 73 6.4 Translation in Eukaryotes 73 6.4.1 Initiation of translation 73 6.4.2 Translation from bicistronic mRNAs 75 6.4.3 Co- and post-translational modifi cation of proteins 77 6.5 Transport of Molecules in Eukaryotic Cells 77 6.6 Transcription and Translation in 80

Chapter 7 Virus Genome Replication 83 7.1 Overview of Virus Genome Replication 84 7.2 Locations of Virus Genome Replication in Eukaryotic Cells 86 7.3 Initiation of Genome Replication 86 7.3.1 RNA and protein primers 86 7.4 Polymerases 88 7.5 DNA Replication 88 7.6 Double-Stranded RNA Replication 90 7.7 Single-Stranded RNA Replication 90 7.8 Reverse Transcription 91

Chapter 8 Assembly and Exit of Virions from Cells 93 8.1 Introduction to Assembly and Exit of Virions from Cells 94 8.2 Nucleocapsid Assembly 95 8.2.1 Helical viruses 95 8.2.2 Icosahedral viruses 95 8.2.3 Genome packaging 96 8.2.4 Assembly mechanisms 96

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8.3 Formation of Virion Membranes 96 8.3.1 Budding through cell membranes 96 8.3.2 De novo synthesis of viral membranes 98 8.4 Virion Exit From the Infected Cell 99

Chapter 9 Outcomes of Infection for the Host 101 9.1 Introduction to Outcomes of Infection for the Host 103 9.2 Factors Affecting Outcomes of Infection 103 9.2.1 Innate in vertebrates 103 9.2.2 Adaptive immunity in vertebrates 105 9.2.3 RNA silencing 108 9.2.4 Programmed cell death 108 9.3 Non-Productive Infections 109 9.3.1 Latent infections 109 9.3.2 Abortive infections 110 9.4 Productive Infections 110 9.4.1 Spread of infections within multicellular hosts 110 9.4.2 Disease 111

Chapter 10 Classifi cation and Nomenclature of Viruses 115 10.1 History of Virus Classifi cation and Nomenclature 116 10.1.1 International Committee on Taxonomy of Viruses 116 10.2 Modern Virus Classifi cation and Nomenclature 116 10.2.1 Classifi cation based on genome sequences 117 10.2.2 Nomenclature of viruses and taxonomic groups 117 10.3 Baltimore Classifi cation of Viruses 119

Chapter 11 Herpesviruses (and Other dsDNA Viruses) 121 11.1 Introduction to Herpesviruses 122 11.2 The Human Herpesviruses 122 11.2.1 viruses 1 and 2 122 11.2.2 Varicella-zoster virus 122 11.2.3 Epstein–Barr virus 122 11.2.4 Human cytomegalovirus 123 11.2.5 Human herpesvirus 6 123 11.2.6 Human herpesvirus 7 123 11.2.7 Kaposi’s sarcoma-associated herpesvirus 123 11.3 The Herpesvirus Virion 123 11.4 HSV-1 Genome Organization 125 11.5 HSV-1 Replication 125 11.5.1 Attachment and entry 125 11.5.2 Transcription and translation 126 11.5.3 Genome replication 126 11.5.4 Assembly and exit of virions from the cell 128 11.5.5 Overview of HSV-1 replication 129 11.6 Latent Herpesvirus Infection 129 11.7 Other dsDNA Viruses 132

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Chapter 12 Parvoviruses (and Other ssDNA Viruses) 135 12.1 Introduction to Parvoviruses 136 12.2 Examples of Parvoviruses 136 12.2.1 Dependoviruses 136 12.2.2 Autonomous parvoviruses 137 12.3 Parvovirus Virion 137 12.3.1 137 12.3.2 Genome 138 12.4 Parvovirus Replication 138 12.4.1 Attachment and entry 138 12.4.2 Single-stranded DNA to double-stranded DNA 139 12.4.3 Transcription and translation 139 12.4.4 DNA replication and virion assembly 140 12.4.5 Overview of parvovirus replication 140 12.4.6 Dependovirus replication 140 12.5 Other ssDNA Viruses 141

Chapter 13 Reoviruses (and Other dsRNA Viruses) 145 13.1 Introduction to Reoviruses 146 13.2 Virion 146 13.3 Rotavirus Replication 147 13.3.1 Attachment and entry 147 13.3.2 Early events 148 13.3.3 Late events 151 13.3.4 Overview of rotavirus replication 152 13.3.5 Rotavirus disease 152 13.4 Other dsRNA Viruses 153

Chapter 14 Picornaviruses (and Other Plus-Strand RNA Viruses) 155 14.1 Introduction to Picornaviruses 156 14.2 Some Important Picornaviruses 156 14.2.1 Hepatitis A virus 156 14.2.2 156 14.2.3 Coxsackieviruses 156 14.2.4 Rhinoviruses 157 14.2.5 Foot and mouth disease virus 157 14.3 Picornavirus Virion 157 14.3.1 Capsid 158 14.3.2 Genome 159 14.4 Picornavirus Replication 160 14.4.1 Attachment 160 14.4.2 Entry 160 14.4.3 Translation and post-translational modifi cations 161 14.4.4 Transcription/genome replication 161 14.4.5 Assembly and exit 164 14.4.6 Inhibition of host gene expression 164 14.4.7 Overview of picornavirus replication 164

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14.5 Picornavirus Recombination 164 14.6 Picornavirus Experimental Systems 165 14.6.1 Reverse genetics 166 14.6.2 Cell-free synthesis of infectious virus 166 14.7 Other Plus-Strand RNA Viruses 167

Chapter 15 Rhabdoviruses (and Other Minus-Strand RNA Viruses) 169 15.1 Introduction to Rhabdoviruses 170 15.2 Some Important Rhabdoviruses 170 15.2.1 Rabies virus 170 15.2.2 Vesicular stomatitis virus 172 15.3 The Rhabdovirus Virion and Genome Organization 172 15.4 Rhabdovirus Replication 172 15.4.1 Attachment and entry 172 15.4.2 Transcription 174 15.4.3 Translation 175 15.4.4 Genome replication and secondary transcription 175 15.4.5 Assembly of virions and exit from the cell 177 15.4.6 Inhibition of host gene expression 177 15.4.7 Role of the nucleus 178 15.4.8 Overview of rhabdovirus replication 178 15.5 Other Minus-Strand RNA Viruses and Viruses with Ambisense Genomes 179 15.6 Reverse Genetics 180

Chapter 16 Infl uenza Virus 183 16.1 Introduction to Infl uenza Viruses 184 16.2 The Infl uenza Virion 184 16.3 Infl uenza A Virus Replication 186 16.3.1 Attachment and entry 186 16.3.2 RNA replication 187 16.3.3 Translation and transport of proteins 188 16.3.4 Virion assembly and release 189 16.3.5 Overview of infl uenza virus replication 190 16.4 Importance of Infl uenza Viruses 190 16.5 Reverse Genetics 192

Chapter 17 Retroviruses 195 17.1 Introduction to Retroviruses 196 17.2 Virion 196 17.3 Retrovirus Replication 196 17.3.1 Attachment and entry 196 17.3.2 Reverse transcription 198 17.3.3 Integration of the provirus 198 17.3.4 Transcription and genome replication 198 17.3.5 Translation and post-translational modifi cations 201

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17.3.6 Assembly and release of virions 202 17.3.7 Overview of retrovirus replication 203 17.4 Examples of Retroviruses 203 17.5 Retroviruses as Gene Vectors 205 17.6 Endogenous Retroviruses 205

Chapter 18 Human Immunodefi ciency Viruses 207 18.1 Introduction to HIV 208 18.2 HIV Virion 208 18.3 HIV Genome 209 18.4 HIV-1 Replication 209 18.4.1 Attachment and entry 209 18.4.2 Reverse transcription and transport to the nucleus 212 18.4.3 Early gene expression 213 18.4.4 Late gene expression 214 18.4.5 Assembly and exit of virions 216 18.4.6 Overview of HIV-1 replication 217 18.5 HIV-1 Variability 217 18.5.1 Antigens 218 18.5.2 Host cell range 218 18.5.3 Resistance to drugs 219 18.6 Progression of HIV Infection 219 18.7 Prevention of HIV Transmission 220

Chapter 19 Hepadnaviruses (and Other Reverse-Transcribing DNA Viruses) 223 19.1 Introduction to Hepadnaviruses 224 19.2 Importance of HBV 224 19.3 HBV Virion 225 19.3.1 DNA 225 19.3.2 P (polymerase) protein 225 19.3.3 Capsid 225 19.3.4 Envelope 226 19.4 Non-Infectious Particles 226 19.5 Soluble Virus Protein 227 19.6 HBV Genome 227 19.7 HBV Genetic Groups 228 19.8 HBV Replication 228 19.8.1 Attachment 228 19.8.2 Entry 228 19.8.3 Transcription 229 19.8.4 Translation 231 19.8.5 Nucleocapsid assembly 231 19.8.6 Genome synthesis 232 19.8.7 Envelope acquisition and exit 232 19.8.8 Overview of HBV replication cycle 232 19.9 Prevention and Treatment of HBV Infection 235 19.10 Other Reverse-Transcribing DNA Viruses 236

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Chapter 20 Bacterial Viruses 237 20.1 Introduction to Bacterial Viruses (Bacteriophages) 238 20.2 Single-Stranded RNA Phages 239 20.2.1 Virion structure of ssRNA phages 239 20.2.2 Genome of ssRNA phages 239 20.2.3 Replication cycle of ssRNA phages 240 20.3 Double-Stranded RNA Phages 243 20.4 Single-Stranded DNA Phages 245 20.4.1 Icosahedral ssDNA phages 245 20.4.2 Filamentous ssDNA phages 247 20.5 Double-Stranded DNA Phages 254 20.5.1 Phage T4 254 20.5.2 Phage T7 256 20.5.3 Phage lambda (λ) 257

Chapter 21 Origins and Evolution of Viruses 263 21.1 Introduction to Origins and Evolution of Viruses 264 21.2 Origins of Viruses 264 21.2.1 Molecular precursors of cellular 264 21.2.2 Components of cells 265 21.2.3 Intracellular micro-organisms 265 21.2.4 How did viruses originate? 265 21.3 Evolution of Viruses 266 21.3.1 Viruses from the past 266 21.3.2 Monitoring virus evolution 266 21.3.3 Mechanisms of virus evolution 267 21.3.4 Evolution of new viruses 273 21.3.5 Co-evolution of viruses and their hosts 273

Chapter 22 Emerging Viruses 277 22.1 Introduction to Emerging Viruses 278 22.2 Viruses in New Host 278 22.2.1 Bunyaviruses 278 22.2.2 Paramyxoviruses 279 22.3 Viruses in New Areas 280 22.3.1 West Nile virus 280 22.3.2 Chikungunya virus 281 22.3.3 Bluetongue virus 282 22.4 Viruses in New Host Species and in New Areas 283 22.4.1 Filoviruses 283 22.4.2 Monkeypox virus 284 22.5 New viruses 285 22.5.1 SARS 285 22.6 Recently Discovered Viruses 285 22.6.1 Human metapneumovirus 285

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22.7 Re-Emerging Viruses 286 22.7.1 and viruses 286 22.7.2 Bioterrorism 286 22.8 Virus Surveillance 286 22.9 Dealing with Outbreaks 286

Chapter 23 Viruses and Cancer 289 23.1 Introduction to Viruses and Cancer 290 23.2 Papillomavirus-Linked Cancers 291 23.3 Polyomavirus-Linked Cancers 291 23.4 Epstein–Barr Virus-Linked Cancers 292 23.5 Kaposi’s Sarcoma 293 23.6 Adult Leukemia 294 23.7 Hepatocellular Carcinoma 294 23.8 Virus-Associated Cancers in Animals 294 23.9 Cell Lines Derived From Virus-Associated Cancers 294 23.10 How Do Viruses Cause Cancer? 295 23.10.1 “Deliberate” interference with control of cell activities 295 23.10.2 “Accidental” activation of cell genes 296 23.10.3 Retroviral oncogenes 296 23.10.4 Damage to immune defenses 297 23.10.5 Overview of cancer induction by viruses 297 23.11 Prevention of Virus-Induced Cancers 299

Chapter 24 Survival of Infectivity 301 24.1 Preservation of Virus Infectivity 302 24.2 Destruction of Virus Infectivity 302 24.3 Inactivation Targets in Virions 302 24.4 Inactivation Kinetics 302 24.5 Agents that Inactivate Virus Infectivity 303 24.5.1 Physical agents 303 24.5.2 Chemical agents 304

Chapter 25 Virus Vaccines 307 25.1 Introduction to Virus Vaccines 308 25.2 Live Attenuated Virus Vaccines 308 25.3 Inactivated Virus Vaccines 310 25.4 Virion Subunit Vaccines 310 25.5 Live Vaccines 310 25.6 Mass-Production of Viruses for Vaccines 311 25.7 Virus-Like Particles 312 25.8 Synthetic Peptide Vaccines 312 25.9 DNA Vaccines 313 25.10 Storage and Transport of Vaccines 313

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Chapter 26 Anti-viral Drugs 315 26.1 Introduction to Anti-viral Drugs 316 26.2 Development of Anti-viral Drugs 316 26.2.1 Screening compounds for anti-viral activity 316 26.2.2 Rational design of anti-viral drugs 316 26.2.3 Safety of anti-viral drugs 317 26.3 Examples of Anti-viral Drugs 317 26.3.1 analogs 317 26.3.2 Non-nucleoside inhibitors of reverse transcription 320 26.3.3 HIV protease inhibitors 320 26.3.4 HIV attachment and fusion inhibitors 321 26.3.5 HIV integrase inhibitors 321 26.3.6 Infl uenza virus neuraminidase inhibitors 321 26.4 Drug Resistance 322 26.5 Anti-viral Drug Research 324

Chapter 27 Prions 327 27.1 Introduction to Prions 328 27.2 Transmissible Spongiform Encephalopathies 328 27.3 The Nature of Prions 328 27.3.1 replication 330 27.4 Prion Diseases 330 27.4.1 Prion diseases in animals 330 27.4.2 Prion diseases in humans 331 27.5 Prion Strains 332 27.6 Prion Transmission 332 27.7 The Protein-Only Hypothesis 332

Virologists’ Vocabulary 335 Index 347

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