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- Brief facts
- Life cycle
- Immune response
- WNV and global warming
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)
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.
HistoryFor decades after its first isolation in 1937 from the blood of a febrile patient in the West Nile Province in Uganda, WNV has been regarded as a minor pathogen. Most of the cases were asymptomatic, and humans and animals were considered as accidental dead-end hosts however sporadic outbreaks of WNV infection associated with encephalitis and death have been reported in Israel (1950s), France (1962-1963), South Africa (1974) and India (1980/1981). Only after the severe human outbreak occurring in Romania in 1996, WNV infection has been declared a major public health and veterinarian concern in Europe and in the Mediterranean basin.
In 1999, WNV unexpectedly emerged in New York, with encephalitis reported in humans and horses. During this outbreak 62 human cases of WNV including 7 deaths were recorded. In the following years, the virus had spread across the United States, Canada, Central and South America. It took only 4 years for it to reach west coast of United States.
RangeWest Nile viruses have a pandemic distribution having been isolated on every continent with the exception of Antarctica. It is the most widespread of the flaviviruses. Migratory birds are thought to be primarily responsible for virus dispersal, including reintroduction of WNV from endemic areas into regions that experience sporadic outbreaks.
Little human WND disease has been reported in Latin America. Several hypotheses were put forward:
- the spread of WNV through migrating birds might select for attenuated viral strains if more virulent strains impair bird migration;
- previous flavivirus infection such as dengue virus might provide some immunity to WNV;
- ecologic conditions in tropical regions might be unfavorable for virulent strains;
- insensitive surveillance and nonspecific laboratory tests could hinder assessment of WNV burden;
- continuous avian host availability for ornithophilic mosquitoes in tropical areas might decrease likelihood of infected mosquitoes biting humans.
DamageBetween 1999 and 2010, ~1.8 million people were infected, with ~360,000 illnesses, 12,852 reported cases of encephalitis/meningitis, and 1,308 deaths. The threat of WNV infection has led to the costly implementation of national blood donor screening, vaccine and drug development, and anti-mosquito spraying. Public outreach campaigns have altered human behavior, including shortening of the time spent outdoors, especially by older people, who are at high risk for WNV disease, and heavy usage of repellents.
The impact of WNV on wild and urban bird populations have been even more severe. Millions of birds have died from WNV infection, and regional-scale population declines of >50% have been observed for several species. The wide range of taxa that have declined includes corvids, chickadees and titmice, wrens, and thrushes and probably other Passeriformes (song birds). Some populations have recovered, whereas others have not.
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.
Since the mid-1990s, three epidemiologic trends have emerged regarding WNV:
- increased frequency of outbreaks in humans and horses,
- increase in reported cases of neuroinvasive disease in humans,
- high fatality rates in birds coinciding with human outbreaks, mainly in the USA and Israel.
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:
- survival of previously infected and now immune birds increases the likelihood of infected mosquitoes feeding on immune hosts;
- mortality in birds is associated with high peripheral viremia leading to the efficient infection of moderately susceptible mosquito vectors;
- immobility of dying bird makes it an easy target whereas hyperthermia increases attractiveness.
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.
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.
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.
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.
The life cycle of WNV within cells is similar to other RNA viruses that replicate cytoplasmically.
- Attachment/entry WNV enters cells via receptor-mediated endocytosis, and is transported into endosomes. The WNV receptor is unknown. Also, the receptor(s) required for WNV binding and entry may vary by cell type. After binding to the cell's membrane, the virus is taken up via clathrin-mediated endocytosis; in the acidified endosome the E protein undergoes conformational changes resulting in fusion between the viral and cellular membranes and release of the viral nucleocapsid into the cytoplasm. Besides the mildly acidic endosomal pH, it is the composition of the target membrane that plays an important role in the membrane fusion process of flaviviruses. For example, in vitro studies have found that addition of cholesterol to target membranes has strong promoting effect on the membrane fusion capacity with TBEV and WNV.
- Translation The viral RNA serves as mRNA and translated into a single polyprotein, which is proteolytically processed to yield three structural proteins (the envelope protein E; the membrane precursor protein prM; and the capsid protein C) and seven non-structural (NS) proteins (NS1, NS2a, NS2b, NS3, NS4a, NS4b, and NS5). The cleavages at the junctions C-prM, prM-E, E-NS1, NS4A-NS4B, and likely also NS1- NS2A, are performed by the host signal peptidase located within the lumen of the ER. The remaining peptide bonds are cleaved by the virus encoded NS3 protease.
- Replication Genome replication is carried out in structures termed replication complexes (RC) that are established by the viral proteins and surrounded host membranes. Replication requires the viral protein NS5, which is an RNA-dependent RNA polymerase. An "antisense" negative strand RNA is produced by this enzyme, which then serves as a template for the synthesis of many new copies of the infectious positive strand of RNA genome.
- Assembly/egress WNV assembles on virus-induced membranes derived from the endoplasmic reticulum and buds into the lumen as immature virions on which E and prM proteins form 60 heterotrimeric spikes. Transit of the immature virion through the mildly acidic compartments of the trans-Golgi network triggers a rearrangement of E proteins on the immature virion; the lower pH induces a structural transition such that E proteins lie flat as antiparallel dimers on the surface of the virion. Under acidic conditions, prM remains associated with the virion and protrudes from the surface of an otherwise smooth virus particle. This pH-dependent conformational change increases the susceptibility of prM for a furin-like serine protease. Cleavage and release of prM completes the virion maturation process, and is a required step in the virus lifecycle. Mature virions exit cells into extracellular milieu by exocytosis and/or budding.
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).
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.
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.
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
- Intrauterine (first documented in 2002 when a woman with WNV encephalitis delivered infected infant with birth defects);
- Infected blood transfusion and organ transplant;
- Needle-stick injuries, inhalation and conjuctival exposure while handling infected birds (laboratories, farms);
- Ingestion of infected prey (alligators, cats, dogs);
- Exposure (potentially) to contaminated feces (American crow can shed >108.8 pfu g-1 in their feces).
Pfeffer M, Dobler G. Emergence of zoonotic arboviruses by animal trade and migration. Parasit Vectors. 2010 Apr 8;3(1):35.
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.
|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|
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:
- disruption of blood-brain barrier (BBB) - upregulation of TNF- α (tumor necrosis factor α) may result in capillary leakage and increased BBB permeability;
- infection or passive transport through the endothelium or choroid plexus epithelial cells;
- infection of olfactory neurons and spread to the olfactory bulb;
- "Trojan horse" mechanism in which the virus is transported by infected immune cells trafficking to the CNS;
- direct axonal retrograde transport from infected peripheral neurons.
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.
|Organism||Incubation 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|
|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.|
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.
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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