Viral Encephalitis

1. Viral Encephalitis Dan Karlin, Jenny Richmond, Chiemi Suzuki BIO 4158: Microbiology and Bioterrorism Dr. Zubay April 20, 2004

2. Roadmap • Introduction • History and epidemiology • Molecular biology • Weaponization • Clinical manifestations • Preparednes and continued surveillance

3. Introduction • Encephalitis is an acute inflammatory process affecting the brain • Viral infection is the most common and important cause, with over 100 viruses implicated worldwide • Symptoms • Fever • Headache • Behavioral changes • Altered level of consciousness • Focal neurologic deficits • Seizures • Incidence of 3.5-7.4 per 100,000 persons per year

4. Causes of Viral Encephalitis • Herpes viruses – HSV-1, HSV-2, varicella zoster virus, cytomegalovirus, Epstein-Barr virus, human herpes virus 6 • Adenoviruses • Influenza A • Enteroviruses, poliovirus • Measles, mumps, and rubella viruses • Rabies • Arboviruses – examples: Japanese encephalitis; St. Louis encephalitis virus; West Nile encephalitis virus; Eastern, Western and Venzuelan equine encephalitis virus; tick borne encephalitis virus • Bunyaviruses – examples: La Crosse strain of California virus • Reoviruses – example: Colorado tick fever virus • Arenaviruses – example: lymphocytic choriomeningitis virus

5. What Is An Arbovirus? • Arboviruses = arthropod-borne viruses • Arboviruses are maintained in nature through biological transmission between susceptible vertebrate hosts by blood-feeding arthropods • Vertebrate infection occurs when the infected arthropod takes a blood meal

6. http://www.cdc.gov/ncidod/dvbid/arbor/schemat.pdf

7. Major Arboviruses That Cause Encephalitis • Flaviviridae • Japanese encephalitis • St. Louis encephalitis • West Nile • Togaviridae • Eastern equine encephalitis • Western equine encephalitis • Bunyaviridae • La Crosse encephalitis

8. http://www.cdc.gov/ncidod/dvbid/arbor/worldist.pdf

9. West Nile Virus

10. West Nile Virus • Flavivirus • Primary host – wild birds • Principal arthropod vector – mosquitoes • Geographic distribution - Africa, Middle East, Western Asia, Europe, Australia, North America, Central America http://www.walgreens.com/images/library/healthtips/july02/westnilea.jpg

11. History of West Nile Virus • 1937 - West Nile virus isolated from woman in Uganda • 1950s – First recorded epidemics in Israel (1951-1954, 1957) • 1962 – Epidemic in France • 1974 – Epidemic in South Africa. Largest ever West Nile epidemic. • 1996 – Romanian epidemic with features similar to those of the North American outbreak. 500 cases and 50 deaths. • 1999 – Russian outbreak. 40 deaths.

12. West Nile Virus: 1999 New York Outbreak • Crows dying in and around Queens in late summer • 27 deaths among captive birds in the Queens and Bronx zoos • Concomitant human infection of apparent encephalitis in the same area • Outbreak was first attributed to St. Louis encephalitis, but tissue samples from dead crows confirmed that it was West Nile virus • 59 human cases requiring hospitalization, including 7 deaths

13. Spread of West Nile Virus in the US • 2000 – spread throughout New England and Mid-Atlantic regions. • 18 new human cases reported • 2001 – spread throughout the entire eastern half of the US • 64 cases reported, with NY, FL and NJ accounting for 60% • 2002 – spread westward across Great Plains into Western US. Reached California by Labor Day. • By end of 2002 cumulative human cases > 3900, with > 250 deaths • 2003 – US, Canada, Mexico • 9,858 cases reported to CDC, including 262 deaths in 45 states and D.C.

14. West Nile Activity in the US – Reports as of April 7, 2004

15. West Nile Activity in the US – Counties Reporting Cases as of March 24, 2004

16. West Nile Virus 2004:BREAKING NEWS • April 13, 2004 – Ohio may have first 2004 West Nile Case • 79 year old man from Scioto County, OH was admitted April 1 with viral meningitis and encephalitis which rapidly progressed to coma over 2 days. • Initial IgM antibody titers were positive for West Nile virus and he complained of itching from insect bites upon admission • Has been treated with blood-pressure drugs to control over-response by the immune system to West Nile virus, causing brain inflammation. • Previously unresponsive and paralyzed. • Can now open his eyes and shake his head in response to questions, but still cannot talk.

17. St. Louis Encephalitis

18. St. Louis Encephalitis • Flavivirus • Most common mosquito-transmitted human pathogen in the US • Leading cause of epidemic flaviviral encephalitis

19. History of St. Louis Encephalitis • 1933 – virus isolated during St. Louis and Kansas City, MO epidemic • 1940’s – virus spread to Pacific Coast • 1959-1971 – virus spread to Southern Florida • 1974-1977 – last major epidemic. Over 2,500 cases in 35 states. • 1990-1991 – South Florida epidemic. 226 cases and 11 deaths. • 1999 – New Orleans outbreak. 20 reported cases.

20. St. Louis Encephalitis

21. Japanese Encephalitis

22. Japanese Encephalitis • Flavivirus related to St. Louis encephalitis • Most important cause of arboviral encephalitis worldwide, with over 45,000 cases reported annually • Transmitted by culex mosquito, which breeds in rice fields • Mosquitoes become infected by feeding on domestic pigs and wild birds infected with Japanese encephalitis virus. Infected mosquitoes transmit virus to humans and animals during the feeding process.

23. History of Japanese Encephalitis • 1800s – recognized in Japan • 1924 – Japan epidemic. 6125 cases, 3797 deaths • 1935 – virus isolated in brain of Japanese patient who died of encephalitis • 1938 – virus isolated from Culex mosquitoes in Japan • 1948 – Japan outbreak • 1949 – Korea outbreak • 1966 – China outbreak • Today – extremely prevalent in South East Asia. 30,000-50,000 cases reported each year.

24. Distribution of Japanese Encephalitis in Asia, 1970-1998

25. Eastern Equine Encephalitis

26. Eastern Equine Encephalitis • Togavirus • Caused by a virus transmitted to humans and horses by the bite of an infected mosquito. • 200 confirmed cases in the US 1964-present • Average of 4 cases per year • States with largest number of cases – Florida, Georgia, Massachusetts, and New Jersey. • Human cases occur relatively infrequently, largely because the primary transmission cycle takes place in swamp areas where populations tend to be limited.

27. History of Eastern Equine Encephalitis • 1831 – First recognized as a disease in horses. Over 75 horses died in 3 counties in Massachusetts. • 1845-1912 – epizootics in Northeast and Mid-Atlantic regions • 1933 – virus isolated from horse brains • 1938 – association of human disease with epizootics. 30 cases of fatal encephalitis in children living in same area as equine cases. • 1947 – largest recorded outbreak in Louisiana and Texas. 13,344 cases and 11,722 horse deaths

29. Western Equine Encephalitis

30. Western Equine Encephalitis • Togavirus • Mosquito-borne • 639 confirmed cases in the US since 1964 • Important cause of encephalitis in horses and humans in North America, mainly in the Western parts of the US and Canada

31. History of Western Equine Encephalitis • Early 1900’s – epizootics of viral encephalitis in horses described in Argentina • 1912 – 25,000 horses died in Central Plains of US • 1930 – San Joaquin Valley, CA outbreak. 6000 cases in horses. Virus isolated from horse brains • 1938 – virus isolated from brain of a child

33. La Crosse Encephalitis

34. La Crosse Encephalitis • Bunyavirus • On average 75 cases per year reported to the CDC • Most cases occur in children under 16 years old • Zoonotic pathogen that cycles between the daytime biting treehole mosquito, and vertebrate amplifier hosts (chipmunk, tree squirrel) in deciduous forest habitats • Most cases occur in the upper Midwestern state, but recently cases have been reported in the Mid-Atlantic region and the Southeast • 1963 – isolated in La Crosse, WI from the brain of a child who died from encephalitis

36. Summary – Confirmed and Probable Human Cases in the US

37. Molecular Biology of Viruses that can Cause Viral Encephalitis Flaviviridae: West Nile Virus Togaviridae: Eastern and Western Equine Encephalitis Bunyaviridae: La Crosse Virus

38. Flavivirus Japanese Encephalitis Virus St. Louis encephalitis virus West Nile Virus

39. Flavivirus: Virus Classification • Family Flaviviridae • 3 Genera • Flavivirus, Pestivirus, Hepacivirus • Flavivirus - 12 Serogroups • Japanese encephalitis virus serogroup • Includes West Nile Virus (WNV), St. Louis Encephalitis, and others

40. Scanned images of West Nile virus isolated from brain tissue from a crow found in New York.

41. Viral Replication Cycle

42. Genome Structure

43. Viral Genome • Positive Strand RNA Genome • 1 ORF – Genome encodes single polyprotein which is subsequently cleaved • 5’ portion • 3 structural proteins • 3’ portion • 7 non-structural proteins • Genome also includes 5’ and 3’ noncoding regions which have functional importance

44. Secondary structure loops

45. 3’ Stem Loop of Plus Strand • Tertiary interactions of 3’ non-coding region serve to stabilize and compact the 3’ region of the genome and may also create binding sites for cellular and/or viral proteins • Pseudoknots – Formed by interactions between 3’ stem loop and adjacent nucleotides • PK1 May be important for minus strand replication • Interacts with cellular proteins • P104, EF-1α, and p84

46. Conserved Secondary and Tertiary Terminal RNA Structures in Minus Strand • Stem loop structures at 5’ and 3’ ends are conserved across flavivirus species suggesting a functional importance for these groups. • Minus strand stem loops may play a role in facilitating the formation of replication complexes and in releasing newly synthesized minus strands from plus strands. • In addition, its interaction with cellular proteins is important for replication.

47. Viral Proteins: Structural and Non-Structural • Structural Proteins • Capsid (C), Membrane (M), Envelope (E) • The envelope - receptor binding • Dimers of E protein arrange their β sheets in a head to tail formation which lie flat on top of the lipid bilayer. The distal portions of these proteins are anchored in the membrane • Non-Structural Multifunctional Proteins • NS1, NS2A, NS2B, NS3, NS4A, NS4B, NS5 • Many functions of non-structural proteins have yet to be determined

48. Viral Non-Structural Proteins • NS1- may play a role in flavivirus RNA synthesis; it has been shown to be essential for negative strand synthesis • NS2A, NS2B, NS4A, NS4B - may facilitate the assembly of viral replication complexes by an unknown mechanism • NS3: Multifunctional • Proteolytic function upon association with NS2B • RNA triphosphatase function thought to be important for the synthesis of the 5’ cap structure • Helicase and NTPase activity • Its activity may be upregulated through interaction with phosphorylated NS5 • NS5 • RNA dependent RNA polymerase • Methyltransferase domain thought to be required for formation of the 5’ cap

49. Model for Closed-Loop Complex Formation in Flaviviruses

50. Togavirus Eastern Equine Encephalitis Virus Western Equine Encephalitis Virus Venezuelan Equine Encephalitis Virus