Workshop on West Nile virus Berlin, 13th -17th of February, 2012 West Nile epidemiology: global situation and recent outbreaks in Europe

Tamás Bakonyi 1, Norbert Nowotny 2

1 Department of Microbiology and Infectious Diseases, Faculty of Veterinary Science, Szent István University, Budapest, Hungary 2Zoonoses and Emerging Infections Group, Clinical Virology, Department of Pathobiology, University of Veterinary Medicine, Vienna, Austria West Nile epidemiology: global situation and recent outbreaks in Europe

• virology • transmission cycle • epidemiology and distribution – West Nile lineages – the emergence of WNV in the USA – the European scenario West Nile epidemiology: global situation and recent outbreaks in Europe

Virology • West Nile virus (WNV) – Flaviviridae, Flavivirus , mosquito-borne group, • Japanese encephalitis virus serocomplex • +ssRNA genome, ~ 11,000 base • 3 structural and 7 non-structural proteins • stem-loop structures at the UTR regions • icosahedral capsid • enveloped • relatively weak resistance (heat, detergents) • strong antigen – the E glycoprotein is the neutralizing Ag – one serotype – cross-reactions with other flaviviruses West Nile epidemiology: global situation and recent outbreaks in Europe

5’- -3’ C ME NS1 NS2A NS3 NS4A NS5 NS2B NS4B West Nile epidemiology: global situation and recent outbreaks in Europe Transmission cycle • natural host of WNV are wild birds – several species are susceptible • USA: 371 bird species – frequent subclinical infections • varying length of viraemia • varying level of virus titre in the blood (amplifying host) • transient shedding with faeces and body fluids – certain species are more vulnerable • USA: 62% of the WNV positive dead birds were American crows and Blue jays • main vectors are mosquitoes – several species are potential carriers • USA: 62 WNV positive mosquito species reported • >98% of the positive pools are of Culex spp . • in Europe the principal vector is usually Culex pipiens – ornithophillic – overwinters in adult form – gradation in late summer and autumn • the extrinsic period is significantly influenced by the weather ratio infected mosquitoes

days after feeding

disseminated infection in mosquitoes

transmitting mosquitoes

Kilpatrik et al., PloS Path. 2008. 4/6, e10000092 West Nile epidemiology: global situation and recent outbreaks in Europe Transmission cycle • natural host of WNV are wild birds • main vectors are mosquitoes • tick are also potential vectors • several other vertebrates are susceptible hosts – mammals (humans, horses), reptiles (alligators), amphibians (frogs) – incidental hosts – the mosquitoes’ choice – dead-end hosts – viraemia is lower than the mosquito infection threshold • possible non-vectorial transmission – iatrogenic – blood transfusion, organ transplantation – intrauterine, lactogenic – reported in humans – accidental – necropsy, laboratory infections – peroral • scavenger birds – carrion feeding (i.e. craws) • birds of prey (i.e. goshawks) • virus shedding by faeces (common coots?) West Nile epidemiology: global situation and recent outbreaks in Europe West Nile epidemiology: global situation and recent outbreaks in Europe Epidemiology and distribution – first isolation: West Nile district of Uganda, 1937 • from a febrile woman – the most widespread flavivirus • Africa, Europe, Asia, Australia (Kunjin), North and South America • WNV lineages – Two main genetic lineages • lineage 1: word-wide distributed – topotype strain: Eg101 (Egypt, 1951) – three clades (~ geographic distribution) – virulence – variants • lineage 2: limited distribution – topotype strain: B956 (Uganda, 1937) – Sub-Saharan Africa and Madagascar – strain Q3574-5 isolated in 1968 from a migrating Barred Warbler ( Sylvia nisoria ) on Cyprus (Watson et al. 1972) Lanciotti et al. Virology 298, 96-105 (2002) West Nile epidemiology: global situation and recent outbreaks in Europe Epidemiology and distribution • WNV lineages – Two main genetic lineages • lineage 1: word-wide distributed • lineage 2: limited distribution – topotype strain: B956 (Uganda, 1937) – Sub-Saharan Africa and Madagascar – emergence in central Europe (Hungary) in 2004 – emergence in Russia (Volgograd) in 2007 – differences in virulence! – Further genetic lineages • lineage 3: Rabensburg strain – first isolation in the Czech Republic, from Cx. pipiens – reisolation in 1999 ( Cx. pipiens ) and in 2006 ( Ae. rossicus ) – no Rabensburg strain associated cases reported in the area – limited pathogenicity in mice West Nile epidemiology: global situation and recent outbreaks in Europe Epidemiology and distribution • WNV lineages – Two main genetic lineages • lineage 1: word-wide distributed • lineage 2: limited distribution – Further genetic lineages • lineage 3: Rabensburg strain • lineage 4: LEIV-Krnd88-190 – isolated in Russia (Caucasus) in 1998 – isolated from ticks ( Dermacentor marginatus ) – virulence is unknown 1a

Lineage 1

1b

Lineage 2

Putative Lineage 3 Putative Lineage 4 West Nile epidemiology: global situation and recent outbreaks in Europe Epidemiology and distribution • WNV lineages – Two main genetic lineages • lineage 1: word-wide distributed • lineage 2: limited distribution – Further genetic lineages • lineage 3: Rabensburg strain • lineage 4: LEIV-Krnd88-190 (Lvov et al., 2004) • lineage 5: Indian isolates (Bondre et al., 2007) • lineage 6: Malaysian isolate (Scherret et al., 2001) • lineage 7: Spanish isolate (Vazquez et al., 2010) Bondre et al., J. Gen. Virol. 88:875-84 (2007) Vazquez et al., 2010. Emerg Infect Dis., 16.: 549-552. West Nile epidemiology: global situation and recent outbreaks in Europe Epidemiology and distribution • The emergence of WNV in the USA – New York (Queens, Bronx) – 1999 • encephalitis in humans, horses, dogs and cats • wild bird mortality • identification of the agent – WNV lin. 1 • genetic characterisation – close relationship with an isolate from Israel (1998) • the route of introduction remained unknown – The virus established itself in the new habitat and quickly spread westwards WNV Neuroinvasive Disease Incidence, by County, US, 1999

N=54

Incidence per million .01-9.99 10-99.99 >=100

Any WNV activity L. Petersen, CDC WNV Neuroinvasive Disease Incidence, by County, US, 2000

N=19

Incidence per million .01-9.99 10-99.99 >=100

Any WNV activity L. Petersen, CDC WNV Neuroinvasive Disease Incidence, by County, US, 2001

N=64

Incidence per million .01-9.99 10-99.99 >=100

Any WNV activity L. Petersen, CDC WNV Neuroinvasive Disease Incidence, by County, US, 2002

N=2946

Incidence per million .01-9.99 10-99.99 >=100

Any WNV activity L. Petersen, CDC WNV Neuroinvasive Disease Incidence, by County, US, 2003

N=2866

Incidence per million .01-9.99 10-99.99 >=100

Any WNV activity L. Petersen, CDC WNV Neuroinvasive Disease Incidence, by County, US, 2004

N=1142

Incidence per million .01-9.99 10-99.99 >=100

Any WNV activity L. Petersen, CDC WNV Neuroinvasive Disease Incidence, by County, US, 2005

N=1294

Incidence per million .01-9.99 10-99.99 >=100

Any WNV activity L. Petersen, CDC WNV Neuroinvasive Disease Incidence, by County, US, 2006

N=1495

Incidence per million .01-9.99 10-99.99 >=100

Any WNV activity L. Petersen, CDC West Nile Virus in Latin America

2003

2002 2004 2002

2002 2004

2004 •2003 2001 2002 2002

2003

2003 2004 2004

L. Petersen, CDC 2006 Argentina Human WNV Disease Cases by Week of Onset, United States, 2006*

600 550 500 450 400 350 300 250

# cases # 200 150 100 50 0 r l l t t n n b b r r r r y y n n g g p p p v v c c a e e a a p p p a a u u e e e c c o o e e a u u J J u u J J F F M M A A A M M J J - - A S S S O O N N D D ------A ------1 8 8 1 5 9 8 2 - 4 8 7 4 4 3 7 0 5 2 5 9 2 6 0 1 5 9 3 2 1 1 1 2 1 2 1 2 1 1 3 1 2 1 2 2 Week ending * Reported as of 5/2/2007, courtesy L. Petersen Reported Human West Nile Virus Cases by Date of Symptom Onset; and Date of First Positive Surveillance Event, Colorado, 2003 110

100

90 human 80 Fever chicken 70 (n=2323) Neuroinvasive 60 mosquito (n=621) 50

No. of cases 40 horse 30

20 bird 10

0

3 3 3 3 3 3 3 3 3 3 3 -0 -0 -0 -0 -0 -0 -0 -0 -0 -0 -0 n n l l g g g t t t t ú ú jú jú u u u p p k k -j -j - - a a a e e -o -o 4 8 - - - z z 0 4 6 0 0 1 1 5 9 -s -s 1 2 0 2 0 1 2 2 6 Symptom onset1 date 2 Source: John Pape, CO DOH Equine and Human WNV Neuroinvasive Disease Cases, United States, 1999-2006

16000 4000

Total Equine: 24,213 cases 14000 Vaccine introduced 3500

12000 3000 HumanCase Reports Total Human: 9,906 cases 10000 2500

8000 2000

6000 1500 Equine Case Reports Case Equine

4000 1000 Equine Human 2000 500

0 0 1999 2000 2001 2002 2003 2004 2005 2006 Reported WNV Disease Cases in Humans, United States, 1999-2006* Other Total Total Year Fever clinical/ Deaths WNND cases Unspec 1999 59 3 0 62 7 2000 19 2 0 21 2 2001 64 2 0 66 9 2002 2,946 1,160 50 4,156 284 2003 2,866 6,830 166 9,862 264 2004 1,148 1,269 128 2,539 100 2005 1,309 1,607 99 3,000 119 2006 1,495 2,616 194 4,269 177 Total 9,906 13,489 637 23,975 962

* Reported as of 5/2/2007, courtesy L. Petersen Clinical manifestation of WNV in the USA

• Asymptomatic (approx. 75%) • West Nile fever (approx. 25%) • Neuroinvasive disease (approx. 1/140) – Encephalitis – Meningitis – Flaccid paralysis

Courtesy L. Petersen, CDC The Hidden Epidemic WNV Infections (n=1.4 million), 1999-2006

Not reported Reported

Fever Fever 322,800 13,484

Asymptomatic Neuroinvasive, Non-fatal 1,040,300 8,942

Fatal, 962

Courtesy L. Petersen, CDC WNV epidemiology and distribution The European scenario • First detection: Hubalek & Halouzka, 1999 – Albania 1958 (serology) • Virus isolation: – France, Russia, Portugal Slovakia, Moldavia, Ukraine, Hungary, Romania, Czech Rep., Italy

virusisolation case • Cases, outbreaks: seropositivity – 1960’s: France, Russia, Spain, Romania – 1970-90’s: Byelorussia, Ukraine, Romania, Czech Republic Zeller & Schuffenecker, 2004 • 2003, France: horse, magpie • 2002-2007, Spain: imperial eagle ( Aquila adalberti ) • 2006-2007, Germany, Poland: seropositive birds Rather widespread WNV outbreak in summer 2008 in northern Italy

G. Savini et al., 2008, Eurosurveillance G. Savini et al., 2008, Eurosurveillance WNV in Hungary

• Serological studies (1969): wild geese, humans • Isolation (1970’s): bank vole, yellow-necked mouse → no documented clinical case • 2003: – Goose encephalitis • Great Plain, middle of August • 6 week-old goslings, 14% mortality • seroconversion, PCR positivity • Lineage 1 WNV strain – Sporadic human meningitis, encephalitis • Great Plain, end of summer – fall • seroconversion • 2004-2005: – encephalitis cases in birds of prey • Körös-Maros National Park • goshawk, sparrow-hawk; CNS symptoms, mortality • Lineage 2 WNV strain! – sheep encephalitis, death • the virus is > 99% identical with the goshawk strain • 2007: – mortality in geese – horse encephalitis – mortality in birds of prey (goshawk, red-footed falcon) – human cases

• 2008 – wild bird, hhorse and human cases – significant geographic sperad – emergence in Austria • 2009 – 2011 – WNV activity in the affected areas in every epidemic season WNV-cases 2004 WNV-cases 2004 -2005 WNV-cases 2004 -2005 -2007 WNV-cases 2004 -2005-2007 -2008 WNV-cases 2004 -2005 -2007 -2008 -2009 WNV-cases 2004 -2005 -2007 -2008 -2009 -2010 WNV-cases 2004 -2005 -2007 -2008 -2009 -2010 -2011 WNV situation in 2010 in Europe WNV situation in 2010 in Europe

• northern Greece – August - October 2010 – a widespread WNV outbreak involving mainly humans – approx. 250 human cases, 191 neuroinvasive, 32 lethal – Whole genome sequencing revealed the Hungarian / Austrian lineage 2 WNV causing the outbreak in Greece – however, with a possibly important aa mutation • Russia – Volgograd region – in summer / autumn 2010 – a widespread WNV outbreak with ~ 231 reported human cases and 6 deaths – aetiological virus seems to be identical to the one, which caused the 2007 Volgograd outbreak – a lineage 2 virus, different from the Hungarian/Austrian/Greek one Papa et al., Clin. Microbiol. Infect., in press

Histidine → Proline WNV situation in 2010 in Europe

• northern Greece • Russia – Volgograd region • Romania – ~ 50 reported human cases – in 2 human samples a lineage 2 WNV is diagnosed – practically identical with the Volgograd strain • Serbia (Novi Sad) – WNV detection in Cx. pipiens pools – > 99% identity with the Hungarian / Austrian / Greek strain Human West Nile Disease cases, Romania 2010 [Sirbu et al., Eurosurveillance 16 (2), 13 January 2011] WNV situation in 2011 in Europe

• Ancona, North-East Italy – WNV detection in a febrile patient – partial sequence identification – > 99% similarity to the Hungarian / Austrian / Greek lineage 2 strain

West Nile epidemiology: global situation and recent outbreaks in Europe

Conclusions

• WNV is a significant human and animal pathogen • increased WNV activity was observed in Europe since 2008 • exotic (lineage 2) strains emerged and are spreading in the continent • endemic (lineage 1 and 3) strains are also present • the virus activity is influenced by the weather • the circulating strains differ in neurovirulence → Diagnostic difficulties → Necessity of monitoring and surveillance activity Acknowledgements:

Jolanta Kolodziejek Zoonoses and Emerging Infections Group, Clinical Virology, Department Helga Lussy of Pathobiology, University of Veterinary Medicine, Vienna, Austria

Bernhard Seidel Office of Ecology Research and Landscape Assessment, Persenbeug, Austria Éva Ivanics Károly Erdélyi Central Veterinary Institute, Budapest, Hungary Krisztina Ursu

Emöke Ferenczi National Center for Epidemiology, Budapest, Hungary

Orsolya Kutasi Large Animal Clinic, Faculty of Veterinary Science, Szent István University, Üll ő, Hungary

Herbert Weissenböck Institute of Pathology and Forensic Veterinary Medicine, Department of Pathobiology, University of Veterinary Medicine, Vienna, Austria

Katharina Brugger Institute of Veterinary Public Health, University of Veterinary Medicine, Vienna, Austria

EDENext and Vectorie EU FP7 projects