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West Nile virus

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Definition

A species of FLAVIVIRUS, one of the Japanese encephalitis virus group (ENCEPHALITIS VIRUSES, JAPANESE). It can infect birds and mammals. In humans, it is seen most frequently in Africa, Asia, and Europe presenting as a silent infection or undifferentiated fever (WEST NILE FEVER). The virus appeared in North America for the first time in 1999. It is transmitted mainly by Culex spp mosquitoes which feed primarily on birds, but it can also be carried by the Asian Tiger mosquito, Aedes albopictus, which feeds mainly on mammals. (MeSH, Year introduced: 1973)

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Taxonomy

viruses - ssRNA viruses - ssRNA positive-strand viruses, no DNA stage - Flaviviridae - Flavivirus - Japanese encephalitis virus group - West Nile virus -

Acronym: WNV

West Nile virus (WNV) is ssRNA(+) virus that belongs to the Flaviviridae family, a large family with 3 main genera (Flavivirus, Hepacivirus and Pestivirus). More than 70 species have been classified in the genus Flavivirus and the majority of these are ARthropod-BOrn viruses (arboviruses).
List of pathogenic ssRNA(+) viruses at MetaPathogen

Serological cross reactivity, nucleotide sequence data, and vector association have grouped WNV within the Japanese encephalitis (JE) virus serocomplex. In addition to WNV, the JE serocomplex includes three other viral species responsible for human disease including Japanese encephalitis (JEV), St. Louis encephalitis (SLEV) and Murray Valley encephalitis (MVEV) viruses.

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Brief facts

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Phylogeny

It has been recently proposed that WNV can be grouped into 7 lineages. Two major genetic lineages diverging by 25 to 30% nucleotides in 255-bp region of the E glycoprotein gene have been well described. Lineage 1 is widespread that can be further subdivided into at least three more clades contains isolates from Europe, the USA, the Middle East, India, Africa and Australia.
Lineage 2 contains isolates from Southern Africa and Madagascar. Since 2004 lineage 2 has also been observed in central and Eastern Europe. In general the lineage 1 viruses are considered to be more virulent than the lineage 2 viruses. However, animal experiments have demonstrated that highly and less neuroinvasive phenotypes exist in both lineages.

The American WNV strain NY99 that caused the outbreaks in USA in 1999-early 2000s might be derivative of a highly neuroinvasive Israeli strain, which was circulating in Israel in the previous year.

The pathway by which WNV reached North America in 1999 remains unknown but several possibilities have been suggested, including mosquitoes being transported by ships and airplanes and/or migratory birds or birds in trade.

For two years, a homogenous viral population (genotype NY99) prevailed in New York State before introduction of a new genotype (WN02) in 2002 containing two non-coding changes in the E (C2466U) and NS5 (C9352U) gene and one coding change in the E gene (U1441C, V159A). WN02 soon became the dominant genotype in the USA, displacing its predecessor by 2004. This displacement was a result of both earlier and more efficient transmission in Culex ssp mosquitoes and increased adaptation to replication at higher temperatures by WN02.

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Evolution

Trends

Since the mid-1990s, three epidemiologic trends have emerged regarding WNV:

Increased virulence as a factor for increased transmission

Although in many cases co-evolution of host and pathogen leads to diminished virulence of the pathogen there may be evolutionary selective pressure for WNV to kill its avian hosts.

High avian mortality is postulated to potentially increase transmission in three ways:

In addition, in contrast to an assumption made in many models of the evolution of virulence, host death from WNV does not appear to reduce the length of the infectious period of the host: most birds that survive WNV infection clear virus from their blood between days 4th and 6th after infection, and most individuals that die from WNV infection do so at approximately the same time. A key question is whether increased replication and virulence of the virus in the avian host would be also damaging to its arthropod vector.

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WNV genome

West Nile virus genome structure
De Filette M et al. Recent progress in West Nile virus diagnosis and vaccination. Vet Res. 2012 Mar 1;43(1):16.

The RNA genome carries 5' cap at the 5' end, and lacks a polyadenylated tail at 3' end. The genomic RNA corresponds to the messenger RNA for the translation of a single long open reading frame (ORF)vinto one large polyprotein that is processed co- and post-translationally, by virally encoded serine protease and multiple host proteinases, into three viral structural proteins (C, pre-M and E) and seven non-structural (NS) proteins (NS1, NS2a, NS2b, NS3, NS4a, NS4b, NS5). Surrounding the ORF are 5' and 3' non coding regions (NCRs) of around 100 nucleotides (nt) and 400-700 nt respectively.

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WNV structure

West Nile virion structure
Petersen LR, Roehrig JT. West Nile virus: a reemerging global pathogen. Emerg Infect Dis. 2001 Jul-Aug;7(4):611-4.

WNV is a spherical particle of approximately 50 nm in diameter: the lipid host cell-derived bilayer membrane (envelope), surrounds icosahedral nucleocapsid core (30- to 35-nm) containing a single stranded positive polarity (sense) RNA genome of about 11 kilobases. The core is composed of multiple copies of a 12-kDa capsid protein (C). The envelope is modified by the insertion of two integral membrane glycoproteins: 180 copies of the envelope protein (E) arranged as homodimers (head-to-tail) and membrane protein prM (in immature virions). Late in virus maturation, the prM protein is cleaved to M protein (8 kDa) by a cellular protease, and the M protein is incorporated into the mature virion's evelope.

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WNV life cycle

West Nile virus life cycle
Copyright 2002 National Academy of Sciences, U.S.A. For non-commercial and educational use. Samuel CE. Host genetic variability and West Nile virus susceptibility. Proc Natl Acad Sci U S A. 2002 Sep 3;99(18):11555-7.

Stages

The life cycle of WNV within cells is similar to other RNA viruses that replicate cytoplasmically.

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Maintenance and transmission

WNV is maintained in nature in an enzootic (non-human equivalent of endemic - characteristic of a certain area) cycle between birds and ornithophilic (feed almost exclusively on avian blood) mosquitoes (predominantly Culex spp).

Mosquito vectors

WNV infection was detected in >60 species (~12 genera including Aedes and Anopheles) of mosquitoes and in some soft and hard ticks (ticks' role in WNV transmission is unclear).

Mosquito species that participate in mosquito-bird-mosquito cycle are referred to as amplification vectors. Mosquito species that feed indiscriminately can transmit WNV to human, horses and other non-avian vertebrates are known as bridging vectors (for example, Aedes vexans, Aedes albopictus).

It was shown that vectors that earlier were considered exclusively ornithophilic can serve as a bridge. One study indicated that C. pipiens mosquitoes in the northeastern USA shift their feeding behavior from highly competent American robins to mammals and humans in the late summer to early fall, coinciding with the emigration of this avian species. This host switching has also been observed in C. tarsalis in the western USA and in C. nigripalpus in Florida.

The principal amplification vectors of WNV: C. pipiens, C. univittatus and C. antennatus in Africa; C. vishnui, C. triaeniorhynchus and C. pseudovishnui in Asia; C. annulirostis in Australia. In USA: C. pipiens, C. restuans and C. salinarius in the northeast, C. tarsalis in the west, C. quinquefasciatus in the south, and C. nigripalpus in Florida.

Many Culex mosquitoes can transmit WNV vertically - from parent through eggs and larva to offspring. It may be one of most important mechanism of WNV persistence in mosquito population.

Culex pipiens, house mosquito at MetaPathogen

Birds

Birds are natural amplification hosts (or reservoir hosts) for WNV. Generally, an infected host must produce a viremia >105 pfu ml-1. WNV has been detected in more than 300 bird species. Passeriformes (song birds) are considered to be the principal reservoir hosts, although competent birds have also been identified in several other orders.

The clinical outcome of infection varies: chickens and turkeys are resistant to disease while crows, Carolina chickadees, tufted titmice, blue jays, American robins, and eastern bluebirds and other Passeriformes are very susceptible. House sparrows develop viremias that exceed 1010 pfu ml-1. American crows exhibit up to 100% mortality and overall American crow population in USA has declined by an estimated 45% since the introduction of the virus in 1999.

Examples of avian reservoir hosts in United States: American robin (Turdus migratorius), house sparrow (Passer domesticus), European starling (Sturnus vulgaris), and crow (Corvus spp).

Ducks (Anseriformes), pigeons (Columbiformes) and woodpeckers (Piciformes) usually generate viremias insufficient to infect mosquitoes.

Turdus migratorius, American robin at GeoChemBio

Other hosts

WNV was detected in >30 species of mammals (alpaca, baboon, bat, black and brown bears, camel, pig, mouse, squirrels, chipmunks, deer, etc.) as well as in alligators and some frogs.

Majority of non-avian vertebrates that are not favored by ornithophilic mosquitoes are incidental (or dead-end) hosts because they usually (but not always), produce insufficient viremia. However, it was shown that golden hamsters, eastern cottontail rabbits, eastern chipmunks, fox squirrel and even alligators can develop WNV viremia exceeding >105 pfu ml-1.

Non-vector-borne transmission modes
West Nile virus transmission cycle

Pfeffer M, Dobler G. Emergence of zoonotic arboviruses by animal trade and migration. Parasit Vectors. 2010 Apr 8;3(1):35.

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Diagnosis

An initial physical examination of patient presented with fever, headache, myalgia, or the more severe symptoms such as meningitis and flaccid paralysis manifested after exposure to mosquitoes suggest WNV infection especially in endemic areas during the summer months.

To confirm the initial diagnosis, specific laboratory tests must be ordered.

TestPositive results
CBC (Complete Blood Count) Anemia, lymphopenia, thrombocytopenia
IgM-antigen-specific ELISA (enzyme-linked immunosorbent assay) WNV-specific antibodies detected
PRNT (Plaque Reduction Neutralization Test) Known virus stock growth inhibited in tissue culture by serum, indicating neutralizing antibodies
NAT (nucleic acid test) PCR amplification directly shows the presence of WNV genome RNA (mutations can impair this test)
Virus isolation/Plaque assay Serum or CSF (cerebrospinal fluid) contain virus as seen in plaque assay
CSF analysis Antibodies and/or virus present by ELISA or plaque assay; elevated protein and increased polymorphonuclear cells; negative Gram stain
EMG/NCS (Electromyogram and Nerve Conduction Studies) Severe effects on anterior horn cells

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Pathogenesis

After mosquito deposited its WNV-contaminated saliva directly into the blood (during feeding) or into skin tissue (during probing), virus is presumed either to spread by blood or to infect resident dendritic Langerhans cells, which then traffic to the draining lymph node. Shortly thereafter, virus amplifies in the tissues and results in a transient, low-level viremia that lasts a few days and typically wanes with the production of anti-WNV IgM antibodies. Following viremia, the virus infects multiple organs in the body of the host, including the spleen, liver, and kidneys. The virus also can enter central nervous system (neuroinvasion). The envelope (E) glycoprotein of WNV has been implicated in neuroinvasiveness. Several mechanisms have been proposed for WNV entry into the CNS:

Retrospective serological studies have indicated that ~80% of WNV infections are asymptomatic. West Nile fever (WNF) is the most common clinical manifestation and is characterized by the development of high fever, chills, rash, headache, myalgia and nausea (non-specific symptoms that cannot be distinguished from other infectious diseases on clinical examination). Symptoms typically abate within 3–5 days of onset and result in lifetime immunity. WNV infection also may result in the development of severe West Nile neuroinvasive disease (WNND) that can be classified into three main clinical syndromes: encephalitis (inflammation of the brain), meningitis (inflammation of the coverings of the brain and/or spinal cord), and acute flaccid paralysis (ATF) (weakness or loss of muscle tone) of the limbs and, in rare cases, of respiratory muscles. ATF is the most distinctive as well as best characterized symptom occurring in up to 50% of patients, sometimes in the absence of other symptoms. The host's vigorous immune response to infection may also contribute to WNV pathogenesis.

Viral entry into the central nervous system (CS) and development of WNND is identified at or near the time of clearance of virus from the peripheral circulation.

Up to 70-75% of survivors of WNND retain permanent neurological sequelae.

OrganismIncubation period (days) Symptoms (%)Duration of clinical signs (days) WNND among all infected (%) Mortality in patients with WNND (%) Comment
Human 2-14 (days) ~20 2-5 ~0.5-1 ~10 Fatigue can persist for over a month; WNND is more common in elderly and immunocompromized
Horse unknown ~8 21 ~8 23-43 Lesions are rarely detected in extraneural tissues; vaccination can reduce risk of death by 44%
Birds (susceptible) 1-3 ~100 1-3 ~100 ~50-100 Lesions in multiple tissues

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Immune response

TypeMechanism
Innate Pathogen recognition receptors (PRRs), such as toll-like receptors (TLRs), detect foreign dsRNA and upon binding, induce secretion of class I interferons (alpha and beta) that are important for limiting virus levels, reducing neuronal death, and increasing survival via activation of various signal transduction pathways.
Humoral The humoral response is characterized by an early occurrence of IgM-dependent neutralizing antibodies. This correlates with clearance of viremia in serum. In contrast, neutralizing antibodies IgG appear later (after about 1 week) when peak viremia is already over and WNV has entered CNS. IgG is responsible for maintenance of long-term immunity to WNV. Neutralizing antibodies are mainly directed against E protein.
Cellular Cytotoxic T-lymphocytes (CTLs) recognize WNV antigens presented on MNCI molecules (major histocompatibility complex class I) of infected cells, lyse these cells and secrete inflammatory cytokines. Deficiencies in either CD4+ or CD8+ T cells are both associated with increased susceptibility to WNV.

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Treatment

Currently, the primary course of action is supportive care. There is no FDA-licensed vaccine to combat WN disease in humans, vaccines are only available for use in horses and geese.

Furthermore, two classical antiviral compounds, interferon and ribavirin, showed promising results in vitro but it is unclear if they are effective in patients. Passively transferring anti-WNV immunoglobulin has been shown to be effective in mouse and hamster models and may be helpful in patients.

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WNV and global warming

WNV genotype WN02, which replaced initial NY99 genotype, has been demonstrated to disseminate more rapidly and with greater efficiency at elevated temperatures, indicating the potential importance of temperature as a selective criterium for the emergence of novel WNV genotypes with increased vectorial capacity. The extrinsic incubation period (EIP) for WN02 viruses is up to 4 days shorter than for NY99 viruses. This shortening of EIP could have been the factor by which this genotype of virus has been selected. Analyses of temperature patterns in the USA have demonstrated an association between above-normal temperatures and epidemics of WNV in northern latitudes.

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References

  1. Pesko KN, Ebel GD. West Nile virus population genetics and evolution. Infect Genet Evol. 2012 Mar;12(2):181-90.
  2. Petersen LR, Hayes EB. West Nile virus in the Americas. Med Clin North Am. 2008 Nov;92(6):1307-22, ix. Review.
  3. Ulbert S. West Nile virus: the complex biology of an emerging pathogen. Intervirology. 2011;54(4):171-84.
  4. Blitvich BJ. Transmission dynamics and changing epidemiology of West Nile virus. Anim Health Res Rev. 2008 Jun;9(1):71-86.
  5. Sips GJ, Wilschut J, Smit JM. Neuroinvasive flavivirus infections. Rev Med Virol. 2012 Mar;22(2):69-87.
  6. Gould EA, Solomon T. Pathogenic flaviviruses. Lancet. 2008 Feb 9;371(9611):500-9.
  7. Smit JM et al. Flavivirus cell entry and membrane fusion. Viruses. 2011 Feb;3(2):160-71. Epub 2011 Feb 22.
  8. Hamer GL et al. Host selection by Culex pipiens mosquitoes and West Nile virus amplification. Am J Trop Med Hyg. 2009 Feb;80(2):268-78.
  9. Kilpatrick AM. Globalization, land use, and the invasion of West Nile virus. Science. 2011 Oct 21;334(6054):323-7.
  10. Murray KO, Mertens E, Despres P. West Nile virus and its emergence in the United States of America. Vet Res. 2010 Nov-Dec;41(6):67.
  11. Rossi SL, Ross TM, Evans JD. West Nile virus. Clin Lab Med. 2010 Mar;30(1):47-65.
  12. Brown HE, Childs JE, Diuk-Wasser MA, Fish D. Ecological factors associated with West Nile virus transmission, northeastern United States. Emerg Infect Dis. 2008 Oct;14(10):1539-45.
  13. Monini M, Falcone E, Busani L, Romi R, Ruggeri FM. West nile virus: characteristics of an african virus adapting to the third millennium world. Open Virol J. 2010 Apr 22;4:42-51.
  14. De Filette M, Ulbert S, Diamond M, Sanders NN. Recent progress in West Nile virus diagnosis and vaccination. Vet Res. 2012 Mar 1;43(1):16.
  15. Brault AC. Changing patterns of West Nile virus transmission: altered vector competence and host susceptibility. Vet Res. 2009 Mar-Apr;40(2):43. Epub 2009 May 1.
  16. Lim SM, Koraka P, Osterhaus AD, Martina BE. West Nile virus: immunity and pathogenesis. Viruses. 2011 Jun;3(6):811-28. Epub 2011 Jun 15.
  17. Wang G, Minnis RB, Belant JL, Wax CL. Dry weather induces outbreaks of human West Nile virus infections. BMC Infect Dis. 2010 Feb 24;10:38.
  18. Bolling BG et al. Transmission dynamics of an insect-specific flavivirus in a naturally infected Culex pipiens laboratory colony and effects of co-infection on vector competence for West Nile virus. Virology. 2012 Jun 5;427(2):90-7.

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Further reading

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