Chapter 13, part A

1. Chapter 13, part A Viruses, Viroids, and Prions

2. Viruses • Viruses contain DNA or RNA • And a protein coat • Some are enclosed by an envelope • Some viruses have spikes • Most viruses infect only specific types of cells in one host • Host range is determined by specific host attachment sites and cellular factors

3. Viruses versus cellular organisms Cellular Organisms • complex organization • both DNA and RNA • carry out cell division • some are obligate intracellular parasites Viruses • simple organization • DNA or RNA but not both (one exception) • unable to reproduce outside of living cells • obligate intracellular parasites

4. Viruses Figure 13.1

5. Helical Viruses Figure 13.4a, b

6. Polyhedral Viruses Figure 13.2a, b

7. Complex Viruses Figure 13.5a

8. Viral Taxonomy • Family names end in -viridae • Genus names end in -virus • Viral species: A group of viruses sharing the same genetic information and ecological niche (host). Common names are used for species • Subspecies are designated by a number

9. Viral Taxonomy • Herpesviridae • Herpesvirus • Human herpes virus 1, HHV 2, HHV 3 • Retroviridae • Lentivirus • Human Immunodeficiency Virus 1, HIV 2

10. Growing Viruses • Viruses must be grown in living cells. • Bacteriophages form plaques on a lawn of bacteria. Figure 13.6

11. The Cultivation of Viruses • requires inoculation of appropriate living host

12. Hosts for animal viruses • suitable animals • embryonated eggs • tissue (cell) cultures • monolayers of animal cells • plaques • localized area of cellular destruction and lysis • cytopathic effects • microscopic or macroscopic degenerative changes or abnormalities in host cells and tissues

13. Growing Viruses • Animal viruses may be grown in living animals or in embryonated eggs. Figure 13.7

14. Growing Viruses • Animal and plants viruses may be grown in cell culture. • Continuous cell lines may be maintained indefinitely. Figure 13.8

15. Virus Identification • Cytopathic effects • Serological tests • Detect antibodies against viruses in a patient • Use antibodies to identify viruses in neutralization tests, viral hemagglutination, and Western blot • Nucleic acids • RFLPs • PCR

16. Hosts for bacteriophages • usually cultivated in broth or agar cultures of suitable, young, actively growing bacteria • broth cultures lose turbidity as viruses reproduce • plaques observed on agar cultures

17. Multiplication of Bacteriophages (Lytic Cycle) • Attachment Phage attaches by tail fibers to host cell • Penetration Phage lysozyme opens cell wall, tail sheath contracts to force tail core and DNA into cell • Biosynthesis Production of phage DNA and proteins • Maturation Assembly of phage particles • Release Phage lysozyme breaks cell wall

18. Bacterial cell wall Bacterial chromosome Capsid DNA Capsid Sheath Tail fiber Tail 1 Attachment:Phage attaches to host cell. Base plate Pin Cell wall Plasma membrane 2 Penetration:Phage penetrates host cell and injects its DNA. Sheath contracted Tail core 3 Biosynthesis: DNA and Protein Figure 13.10.1

19. Tail DNA 4 Maturation:Viral components are assembled into virions. Capsid 5 Release:Host cell lyses and new virions are released. Tail fibers Figure 13.10.2

20. One-step Growth Curve Figure 13.11

21. Lytic cycle Phage causes lysis and death of host cell • Lysogenic cycle Prophage DNA incorporated in host DNA

22. The Lysogenic Cycle Figure 13.12

23. Specialized Transduction gal gene Bacterial DNA Prophage 1 Prophage exists in galactose-using host (containing the gal gene). Galactose-positive donor cell gal gene 2 Phage genome excises, carrying with it the adjacent gal gene from the host. 3 Phage matures and cell lyses, releasing phage carrying gal gene. gal gene 4 Phage infects a cell that cannot utilize galactose (lacking gal gene). Galactose-negative recipient cell 5 Along with the prophage, the bacterial gal gene becomes integrated into the new host’s DNA. 6 Lysogenic cell can now metabolize galactose. Galactose-positive recombinant cell Figure 13.13

24. Multiplication of Animal viruses • Attachment Viruses attaches to cell membrane • Penetration By endocytosis or fusion • Uncoating By viral or host enzymes • Biosynthesis Production of nucleic acid and proteins • Maturation Nucleic acid and capsid proteins assemble • Release By budding (enveloped viruses) or rupture

25. Virus Purification and Assays • development of virology closely linked to development of these methods

26. Virus Purification • four commonly used methods • differential centrifugation and density gradient centrifugation • precipitation of viruses • denaturation of contaminants • enzymatic digestion of cell constituents

27. Differential centrifugation • separates based • on size

28. Density gradient centrifugation

29. Measuring concentration of infectious units • plaque assays • dilutions of virus preparation made and plated on lawn of host cells • number of plaques counted • results expressed as plaque-forming units (PFU)

31. Measuring concentration of infectious units… • infectious dose and lethal dose assays • determine smallest amount of virus needed to cause infection or death of 50% of exposed host cells or organisms • results expressed as ID50 or LD50

32. Graph