single strand, negative sense RNA Viruses

1. single strand, negative sense RNA Viruses • Elliot J. Lefkowitz

2. Contact Information: Elliot Lefkowitz, Ph.D.Associate Professor, Microbiology • Email • ElliotL@uab.edu • Web Site • http://www.genome.uab.edu • Office • BBRB 277A • Phone • 934-1946

3. Objectives • To understand the fundamental common and distinguishing properties of (-) ssRNA viruses • To understand the basic replication strategies of (-) ssRNA viruses • To be able to identify human pathogens that belong to (-) ssRNA virus families, and some of their biological and pathogenic properties

4. Reading • Medical Microbiology, Murray et al. 6th Edition • General classification • Chapter 4 • RNA virus properties and replication • Chapters 58, 59, 60, 63 • Pathogenesis • Chapters 48, 67

5. Slide References • Fields Virology, 5th Edition • Viruses and Human Disease • Strauss and Strauss • University of Leicester - Virology Online • http://www-micro.msb.le.ac.uk/3035/index.html • International Committee on Taxonomy of Viruses • The 9th ICTV Report • Primary literature

6. Virus classification

7. The Virus World

8. The (-) RNA Virus World

9. RNA Virus Genome Structure • Number of strands • Single or double stranded • Strand polarity • Positive, negative, or ambisense (both + and -) • Positive (Plus) sense denotes the coding (mRNA) strand • Number of segments • Single or multi-segmented

10. single strand RNA virus genome polarity (+) sense RNA virus virion RNA (+) sensevirus mRNA (+) sense 5’ 3’ translation (-) sense RNA virus 3’ 5’ virion RNA (-) sense transcription 5’ 3’ virus mRNA (+) sense translation

11. Negative/AmbisensessRNA Viruses

12. Properties of (-) sense ssRNA Viruses • Enveloped virion • Helical nucleocapsid • Negative-sense, linear, single segment RNA genome • Bornaviruses, Filoviruses, Rhabdoviruses, Paramyxoviruses • Negative and Ambisense, linear, multi segment RNA genomes • Arenaviruses, Bunyaviruses, Orthomyxoviruses • Cytoplasmic replication • Exception: Bornaviruses, Orthomyxoviruses • Genomes are non-infectious • An initial round of transcription is required for genome replication • Virion must contain proteins required for transcription

13. Order: Mononegavirales:Single segment, (-) sense, ssRNA • Bornaviridae • Bornavirus • Filoviridae • Marburg virus • Ebola virus • Paramyxoviridae • Paramyxovirinae • Henipavirus • Morbillivirus • Respirovirus • Rubulavirus • Pneumovirinae • Pneumovirus • Metapneumovirus • Rhabdoviridae • Vesiculovirus • Lyssavirus

14. Multi-Segment, (-) sense ssRNA viruses • Orthomyxoviridae • Influenzavirus A • 8 genome segments • Influenzavirus B • 8 genome segments • Influenzavirus C • 7 genome segments • Isavirus • 8 genome segments • Thogotavirus • 6 genome segments

15. Multi-Segment, Negative and Ambisense ssRNA viruses • Arenaviridae • Two ambisense RNA segments • Bunyaviridae • Three RNA segments • Both negative-sense and ambisense segments • Depends on genus

16. The Virus Virion, Genome, Proteins

17. Viral Proteins • Attachment/entry • G – Membrane glycoprotein • F – Fusion protein • H – Hemagglutinin • N – Neuraminidase • Structural/Assembly • M – Matrix • Underlies lipid bylayer • Replication • N – nucleocapsid protein • P – Phosphoprotein • L – RNA dependent RNA polymerase

18. Rhabdovirus Virion

19. Virus replication Machinery • Proteins • RNA-dependent RNA-polymerase (RdRp) • Transcription • Replication • Nucleocapsidprotein (N) • Encapsidates RNA • Forms helical nucleocapsid • P protein • Phosphoprotein - polymerase cofactor • Forms complexes with N and L • Binds to RNA termini • RNA Genome

20. Genome Organization Mononegavirales Filoviridae Paramyxoviridae Rhabdoviridae

21. Genome Organization Arenaviridae Bunyaviridae

22. Influenza A Genome Structure

23. Virus Coding Strategies • Individual ORFs • Multiple transcripts with transcription attenuation • Polyprotein processing • Single transcript to Large polyprotein: Proteolytic processing • RNA Editing • Insertion/deletion of additional residues (at a specified site) altering the reading frame • Multiple ribosomal initiation sites • Stop codon read-through

24. Virus Replication

25. RNA-dependent RNA Polymerase(RdRp – L Protein) • Catalytic subunit of the polymerase complex • Polymerization of nucleotides • Transcription of mRNA • Capping • Methylation • Polyadenylation • Genome Replication • Most conserved protein between the mononegavirales virus families

26. Source of the RNA-dependent RNA Polymerase • Host cells do not have a suitable one • Therefore the virus must provide its own • RNA viruses use 2 different strategies to provide the RdRp: • Synthesized immediately upon entry and unpackaging of the virion into the cell (positive-sense viruses) • Therefore protein synthesis is the first step in the replication process • Packaged within the virion (negative-sense viruses) • Therefore mRNA transcription is the first step in the replication process

27. VSV Transcription & Replication

28. (-) sense ssRNA virus Human Pathogens

29. Major Viral Target Tissues

30. Arenaviruses/Bunyaviruses

31. Arenavirus and Bunyavirus Disease • Arenaviruses • Mostly rodent viruses • Human zoonoses • Junin virus • Argentine hemorrhagic fever • Lassa Fever • Bunyaviruses • Large group of arthropod-borne viruses • Human pathogens – hemorrhagic fever • Hantaviruses • Rodent-borne • Pulmonary Syndrome/Hemorrhagic fever • Rift Valley Fever virus • Mosquito-borne virus

32. Filoviruses

33. Filovirus Disease

34. Rhabdoviruses

35. Rhabdoviruses

36. Rabies virus Pathogenesis

37. Paramyxoviruses

38. Paramyxoviruses

39. Human Respiratory Syncytial virus • Major cause of lower respiratory tract infections • Rarely life-threatening • Individuals get repeat infections • Highly infectious • Spread is by exchange of respiratory secretions • Infection confined to respiratory tract • Globally: 100,000,000 infections/year • 200,000 deaths/year • In USA: All infants by age of 4 years are infected • 100,000 hospitalizations/year • Estimated cost of $300,000,000/year (1985) • 25-50% of hospital staff infected during outbreaks

40. Measles virus • Extremely infectious • Spreads through contact with respiratory secretions • Victims are infectious before symptoms are evident • Develops systemic infection • Globally: 45,000,000 infections/year • 1,000,000 deaths/year • In USA: Infections are rare • Occasional epidemic in unvaccinated populations • MMR (Measles, mumps, and rubella) vaccine highly effective (2 shots)

41. Acute Disseminated Encephalomyelitis Measles Inclusion Body Encephalitis Subacute Sclerosing Panencephalitis Neurologic Complications of Measles

42. Orthomyxoviruses

43. Orthomyxoviruses • Influenza • A: Mild to severe disease involving upper and especially lower respiratory tract • B: Similar spectrum of illness to A but generally more mild • C: Sporadic upper respiratory illness in humans • 96% of human adults have antibodies • Thogotovirus • Natural host: Ticks • Also infects: Humans, cattle, goats, waterfowl, etc. • Isavirus • Infectious salmon anemia virus

44. Schematic diagram of influenza A viruses G Neumann et al. Nature 000, 1-9 (2009) doi:10.1038/nature08157

45. M2 Ion Channel • Involved in virion uncoating • Highly conserved • Target for amantadine

46. Hemagglutinin

47. Neuraminidase • Virion release from cell membrane • Cleavage of sialic acid from cell membrane thus preventing binding by HA • Target for Oseltamavir (Tamavir) and Zanamivir (Relenza)

48. Variation and evolution • Influenza virus

49. Influenza Virus Variation • Antigenic drift • Amino acid changes • Antigenic shift • Reassortment/exchange of genome segments between strains • Recombination • Detected but rare

50. Reassortment