West Nile virus (WNV)

Epidemiology : first identified in 1937 in Uganda (B 956 strain), WNV is endemic in localities along the flight paths of migratory birds from South Africa across the Mediterranean Sea to Europe, Caspian Sea and south part of Western Siberia. it has been known to cause asymptomatic infection and fevers in humans in ...

Africa : infection of humans and mammals (horses in particular) throughout the African and Eurasian ranges is sporadic and rare. But significant outbreaks, like the 2005 in Astrakhan, have been recorded previously. An epidemic in Romania involved hundreds of human cases with 17 fatalities, and sporadic WNV cases have been recorded in Israel over several years. Recently outbreaks have occurred in horses in France and Italy. WNV-seropositive birds have been recorded in Poland, the Czech Republic and the UK^ref. Further information would be welcomed concerning any common features associated with the WNF outbreaks in Astrakhan in 1999 and 2005
Europe : the majority of human cases have been reported from the eastern Mediterranean region. In the Old World, WNV causes relatively mild infections with influenzalike symptoms or no apparent disease (Peiris JSM, Amerasinghe FP. West Nile fever. In: Beran GW, Steele JH, editors. Handbook of zoonoses. Section B: Viral. 2nd ed. Boca Raton (FL): CRC Press; 1994. p. 139–48); encephalitis and fatalities in the human population, horses, or poultry are spasmodic^{ref1,
ref2,
ref3}

southern France in the 1960's (after the human and equine outbreak in Camargue in 1962-1965, neither human nor equine cases were detected until 2000 in France. From late August until early November 2000, 131 suspected equine cases/76 confirmed cases equine cases were identified in Camargue^ref. In 2003, 7 human cases (3 encephalitis and 4 cases of febrile illness) and 4 equine cases were detected in the Var (another Mediterranean region of France)^ref. A low level of WNV activity was reported in the Camargue region in sentinel birds: one seroconversion in 2001, one in 2002 and none in 2003. In late July 2004, a WNV seroconversion was reported in a sentinel chicken from Saintes-Maries de la Mer, and a second seroconversion was reported in mid-August at the same location. On 6 Sep 2004, 2/3 of the sentinel birds from this flock were positive for WNV antibodies. A sentinel duck was reported to be positive for WNV on 16 Aug (infection confirmed on 7 Sep 2004) in Saint-Just, Herault. By 30 Sep 2004 (week 40), 37 suspected cases in horses, including 4 fatalities or euthanasia, occurred in Saintes-Maries de la Mer, Camargue^ref1,
ref2)
recent cases in western Europe (Denmark, the Netherlands, and Sweden) are considered to have been the result of infection contracted abroad.
Russia : epidemic in southern Russia in 1999 (> 800 cases); isolated in Volgograd in 1999 and in Siberia in 2002 and 2003 from rooks (Corvus frugilegus) and grey crow (Corvus tristis); 4 positive samples from cinereous vultures (Aegypius monachus) and 2 positive samples from the cattle egret (Bubulcus ibis) were found in dead birds collected in the Far Eastern Region of Russia during 2003-2004. Sequencing of 400 bp fragments of the E protein gene showed high homology with the WNV/LEIV-Vlg99-27889 strain of West Nile virus (isolated in Volgograd, Russia, in 1999). Additionally, 4 positive samples from other species of birds (Ixobrychus eurhytmus, Pica pica, Corvus macrorhynchos, Falco tunnuncules) collected during the aitumn of 2004 were found during screening with anti-West Nile virus MAb-modified ELISA), but human cases are not commonly reported in these regions. In Astrakhan since Sep 2005 there were 73 cases and 3 deaths : the Astrakhan region is located in the extreme southeast of the European part of Russia. The region occupies an area of 44 000 sq km. The Astrakhan region borders the Republic of Kazakhstan, the Republic of Turkmenistan, and the Islamic Republic of Iran. Within the Russian Federation the Astrakhan region borders the Volgograd region and Republic of Kalmykia.Endemic areas for West Nile virus in the former Soviet Union include Ukraine, Belarus, the southern area of European Russia (regions of

since the 1970s, illness has been observed in Kazakhstan and the republics of Central Asia, Astrakhan region (in Russia), Ukraine, and Azerbaijan.
high risk exists in the desert territories of the Volga basin. Seropositive humans have been identified in Armenia and Georgia.
seropositivity rates of 0.4-8% were demonstrated among healthy blood donors in European Russia and western Siberia during 1963 to 1993.
1.7% of the general Belarusian population are seropositive: 5.8% in Gomel, and 15.4% in Brest Region; 0.6 to 5.8% of cattle, 2.9 to 6.8% of wild small mammals, and 6.5 to 16.7 percent of birds.

1963 to 1968 : at least 10 cases of human disease were reported in the Volga Delta. At the time, the virus was recovered from ticks, water birds and mosquitoes.
1977 : cases of human disease were reported in the area of Brest (Belarus). Additional cases were reported during the 1980s.
1997 : 38 cases (16 with encephalitis) were reported in Ukraine.
1999 : an outbreak (826 suspect cases, 84 with meningoencephalitis, 40 fatal) was reported in southern Russia (Volgograd, Astrakhan, and Krasnodar regions). Seropositivity rates in the Astrakhan region increased from 31.6% in 1998 to 44.4% in 1999.
2002 : an outbreak (33 cases) was reported in the Astrakhan region, most from the Volga middle delta.
2004 : 3 cases were reported in West Siberia.
2005 : an outbreak (73 cases, 3 fatal) was reported in the Astrakhan region and 15 in the Rostov region.

Culex pipiens

C. modestus

Aedes vexans

the key ornithophilic vector is Culex molestus. Both C. modestus and C. pipiens feed on birds as well as humans.
additional regional tick vectors include Ornithodoros capensis (Azerbaijan), Hyalomma marginatum (Astrakhan and Azerbaijan), Hy. detritum (Turkmenistan) and Rhipicephalus turanicus (Azerbaijan).
potential anthropophilic mosquito vectors in the Volgograd region include Culex modestus, Aedes vexans, Coquillettidia richiardii, Ae. caspius, and Cx.pipiens.
local vectors in Ukraine include Aedes cantans, Ae. caspius, Ae. circumluteolus, Ae. excrucians and Anopheles maculipennis.In the Volga Delta, 56 species of birds are involved in virus circulation. In the coastal area of the Delta, the most important hosts are shore birds (notably in the order Gressores): the green heron (Nicticorax nicticorax) - of which 45% are seropositive; great cormorant (Phalacrocorax carbo); coot (Fulica atra), water hen (Gallinula chloropus); and great grebe (Podiceps cristatus); and to a lesser extent gulls and terns.
in the agricultural region of the Delta, 20 species of birds (particularly rooks, crows, and pigeons) are involved in virus circulation.
in the Kuban and Terek River Deltas, the most important birds are herons, coots, and some wild ducks.
West Nile virus was identified in the southern portion of Western Siberia during 2002 in dead rooks (Corvus frugilegus), teal (Anas crecca) and garganey (Anas querquedula).
circulation of West Nile virus among birds was confirmed in the far eastern region of Russia during 2003 to 2004 - among cinereous vultures (Aegypius monachus) and cattle egrets (Bubulcus ibis).
during 2004, infection was also confirmed in Ixobrychus eurhytmus, Pica pica, Corvus macrorhynchos and Falco tunnuncules.
wild birds which nest in Azerbaijan are involved in the transmission of West Nile virus in the European region.

Poland, the Czech Republic and the United Kingdom : WNV-seropositive birds have been recorded
Italy : since August 1998 there was an outbreak in horses of West Nile encephalitis in Tuscany (region) specifically in the provinces of Pistoia, Lucca, Pisa and Florence : there were 6 officially reported cases (with several tens not reported, including some outside of Tuscany). Some horses were euthanized but some recovered after treatment. A sanitary cordon was established by the authorities encompassing twenty boroughs within the above mentioned provinces : this cordoned off area includes the Bientina and Fucecchio wetlands where the is an abundance of water fowl and mosquitoes. The cases were reported from horse farms in these areas. The preliminary diagnosis, on the basis of serology, was confirmed by virus isolation at the Pasteur Institute of Paris. There were no new cases for more than a month. No human cases were diagnosed, although about 4% of individuals in close contact with these animals seroconverted : molecular studies showed that the virus RNA had >99% nucleotide identity with other WNV strains isolated in Europe and with African strains isolated from Culex univittatus in Kenya and Culex neavei in Senegal. Presumeably migratory birds brought the virus from Africa to Italy. The most probable vectors were Culex mosquitoes, most likely Culex impudicus in the Fucecchio marshes, where it was the predominant mosquito, and Culex pipiens , which was the most common species in the Cerbale hills : both species are primarily avian feeders, but nevertheless Culex pipiens might act as a bridge vector transmitting the virus from birds to horses, and perhaps to humans. Surprisingly there is no mention of the morphologically indistinguishable Culex pipiens s.s. x Cules pipiens molestus , a mosquito that feeds not on birds but predominantly on mammals, including humans^ref. In the past the presence of a West Nile-related virus had been hypothesized on the basis of serologic survey by HI on sheep and goats, but the presence of true neutralizing antibodies had never been demonstrated, nor any virus from mosquitoes collected in these regions had been isolated.
Romania : an epidemic in Romania involved hundreds of human cases with 17 fatalities. A recent investigation of epidemic WNV infection indicated an apparent-to-inapparent infection ratio of 1:325. The Romanian epidemic took us all by surprise in that antibody to WNV had been found in many human and other vertebrate populations in Europe and WNV had been isolated only a few times in Portugal and France but few human or equid cases had been identified; antibody to WNV was detected in humans and livestock in the former Yugoslavia (Croatia), which is quite near to Italy. Most of the early antibody determinations had been made using the broadly cross-reactive hemagglutination-inhibition test, suggesting that we were dealing with a virus closely related to WNV but not necessarily that virus. Given the Romanian epidemic in humans and these cases in Italian horses, it appears that either WNV infections have been under recognized or ignored, or, at least as likely, that WNV is an emerging disease threat to both humans and equids.
Hungary : until 2003, however, WNV infections in Hungary have never been associated with clinical symptoms, although a severe outbreak of West Nile encephalitis in humans was reported in 1996 and 1997 in neighboring Romania. In late summer 2003, an outbreak of encephalitis emerged in a Hungarian goose flock, resulting in a 14% death rate among 6-week-old geese (Anser anser domesticus). Based on histopathologic alterations, serologic investigations, and nucleic acid detection by RT-PCR, WNV was diagnosed as the cause of the disease (Glávits R, Ferenczi E, Ivanics É, Bakonyi T, Mató T, Zarka P, et al. Occurrence of West Nile Fever in a circovirus infected goose flock in Hungary. Avian Pathol. 2005;34:408–14). Chronologically and geographically related to the outbreak in geese, a serologically confirmed WNV outbreak was also observed in humans, which involved 14 cases of mild encephalitis and meningitis (Ferenczi E, Rácz G, Faludi G, Czeglédi A, Mezey I, Berencsi G. Natural foci of classical and emerging viral zoonoses in Hungary. In: Berencsi G, Khan AS, Halouzka J, editors. Emerging biological threat. Amsterdam: IOS Press; 2005. p. 43–9). The phylogenetic analysis emphasizes the close genetic relationship of the goose-Hungary/03 strain with a WNV strain isolated in Israel in 1998 and the WNV strain introduced in New York in 1999, since the 3 WNVs form 1 single cluster within clade 1a of lineage 1. These strains caused outbreaks in birds, humans, and horses. Previous European WNV isolates exhibited lower identity values, e.g., the strain that was responsible for the Romanian outbreak(s) in 1996 and 1997 showed only 96% nt identity with the Hungarian goose-2003 strain, and in the phylogenetic tree the other European isolates form a separate cluster consisting of 2 subclusters. The earliest representatives of the Israel/USA/goose-Hungary/03 cluster were reported by Malkinson et al.^ref from ill and dead white storks (Ciconia ciconia) in Israel in 1998. These storks, however, had hatched in central Europe, and during their autumn migration southwards, strong winds had blown them off course, from their usual route to Africa, to southern Israel. Malkinson et al. suspected that these birds introduced the neurovirulent genotype of WNV to Israel from their hatching place. The wetlands of southeastern Hungary are foraging and nesting habitats for storks and many other wild bird species, and the goose farm, where the WNV outbreak occurred in 2003, is located in this region. These facts, together with the close phylogenetic relatedness of the Israeli/US/Hungarian WNV strains, strongly support the theory that storks carried the neurovirulent WNV strain from central Europe (that is, from Hungary) to Israel, which sheds new light on the introduction of WNV to New York. This virus could have originated in Israel (which is the generally accepted although not proven theory) or central Europe. In both cases, however, the virus seems to have its true origin in Europe. In a recent publication, Lvov et al. suggested that WNV could have been introduced into New York by ships traveling from Black Sea ports^ref. One year later, in August 2004, a goshawk (Accipiter gentilis) fledgling showed CNS symptoms and died in a national park in southeastern Hungary. When histopathologic methods and RT-PCR were used, WNV antigen and nucleic acid were detected in the organs of the bird. Furthermore, the virus was isolated after injection of suckling mice^ref
Germany : 3 imported cases of WNV infection occurred since 2003 : in 2003, a 77 year old man from Lower Saxony and a 51 year old woman from Bavaria became ill with WNV infection after traveling in areas of high prevalence in the USA^ref, while in Sep 2004 a 77 year old woman from Weimar became ill during a tourist trip to California in the US lasting from 4 Sep to 4 Oct 2004^ref.
UK : evidence of WNV circulation in UK birds, probably introduced by migratory birds from overseas, has been reported from presence of WNV neutralizing antibody and small fragments of RNA with sequences corresponding to WNV^ref. WNV-specific seroconversion in sentinel chickens raised on an English farm were reported in 2006. Maternal neutralizing antibodies to WNV in hatchlings declined within 3 weeks. During the following months, healthy chickens developed WNV neutralizing antibodies that were confirmed by immunoblotting and indirect immunofluorescence tests using WNV antigens. The proportion of seropositive chickens was higher for WNV than for Usutu virus or Sindbis virus. Attempts to isolate infectious virus or to detect viral RNA in the sera, failed^ref. The seroconversions in the absence of human or equine cases is interesting, and suggests that prospective monitoring of humans, animals and mosquitoes for additional evidence of WNV transmission would be worthwhile, and provide isolates of the virus for comparison with other WNV isolates. The question of endemicity of WNV versus reintroduction into the UK remains open.

Western Hemisphere : human and animal infections were not documented in North America until 1999^ref1,
ref2. In the New World, WNV exhibits increased virulence among the local wild bird populations and causes more frequent severe central nervous system symptoms and deaths in humans and horses^ref1,
ref2. Although exactly how WNV was introduced into New York is unclear, phylogenetic comparison of the viral nucleic acid sequences has shown a close relationship between the American WNV isolates and strains isolated from encephalitic geese and storks in Israel in 1998^{ref1,
ref2,
ref3}. Experimental infections of rodents indicated that the neurovirulence of WNV correlates with its genotype, and the North American strains are highly neurovirulent for mice^ref. The earliest representatives of the Israel/USA/goose-Hungary/03 cluster were reported by Malkinson et al. (23) from ill and dead white storks (Ciconia ciconia) in Israel in 1998.

USA : in 1999^ref and 2000, outbreaks of WNV encephalitis were reported in persons living in the New York City metropolitan area, New Jersey, and Connecticut. It is thought to have arrived in the USA from Israel or Egypt. In these 2 years, 83 human cases of West Nile illness were reported and 9 died. Then WNV swept to the west coast (including California) and Central America : in 2001, human infection with WNV occurred in 10 states with 66 cases and 9 deaths; in 2002, WNV activity spread to 44 states, with 4,156 human cases and 284 deaths; in 2003, > 230 deaths in 45 states. With the exception of Alaska and Hawaii (where a wild sparrow found at Kahului Airport tested positive at ELISA test but negative on PCR on Sep 2004), Oregon is the last of the states to confirm isolation of WNV in 2004, which can now be considered to be. It spreads more in USA than in Europe due to the different genotype of Culex pipiens , the existence of other mosquito species that are prominent in the west and south and that bite both birds and humans : the circulating viral strain may also be particularly harsh - and American animals have little natural immunity to it as birds have never seen WNV at all. It occurs chiefly in the summer. WNV is now the predominant circulating arthropod-borne virus in the USA with >15,000 human cases and >600 fatalities since 1999
Mexico^ref
Canada : first cases occurred in 2004
Cuba : first 3 human cases (all recovered) and 4 equine cases occurred in Jan 2005. Antibodies were found in 3 birds ("2 red-legged thrushes and a little blue heron") found during field work conducted in 2004^{ref1,
ref2,
ref3,
ref4}. It's the 1st year they've looked in Cuba, but it's only 90 miles from Miami, Florida, where birds have tested positive for at least 2 years. An outbreak in horses in Havana city was reported in Feb 2005 with a the seroprevalence of 10.6% and lack of deaths or clinical signs. In serosurvey of healthy horses on Guadeloupe in 2002, in 6 months, the prevalence increased from 2.8 to 50%, so if this is a recent outbreak we might expect more horses to seroconvert. It would be important to confirm if there were actually no clinical signs in any of the outbreak horses since the emergence of the disease in Cuba, or if the horses just weren't showing signs at the time of the serosurveillance. The report seems to indicate that there have been no clinical signs. If accurate, then it would imply that a newly emerged West Nile virus caused little or no illness in equines.
Puerto Rico : 2 resident birds, a bananaquit and a green heron, with antibodies to WNV
Argentina^ref : in April 2006 at least 2 Pure Bred racehorses from a ranch located in San Antonio de Areco, in the county of Capitan Sarmiento, in Buenos Aires Province, about 113 km. northwest from the capital city of Buenos Aires died and were confirmed by OIE to be affected by WNV^ref. This is the 1st report of disease due to WNV in South America. ProMED mail's prior report on the most southerly extension of the virus in the Western Hemisphere was in Trinidad with a report of 2 seropositive horses and 3 seropositive birds. One wonders if the other countries along the route^ref have had WNV infections but have not as yet confirmed the presence of the virus in their countries. Information from knowledgeable sources in Argentina, and in other countries in South America on WNV activity in their countries would be very much appreciated. While WNV has not commonly been reported from waterfowl, there has been a wide range of species that have been infected with the virus as seen in the North American experience. Newswires speculated WNV as a possible etiology for the avian die-off, mentioning that it has already entered South America^ref. An excellent review of the North American experience with WNV^ref

South-West Asia

Japan : first cases occurred in 2004; a Japanese man in his 30s who stayed in Los Angeles from 28 Aug to 4 Sep 2005 was found infected with WNV on Mon 3 Oct 2005. This is the 1st serologically confirmed human case of WNV infection in Japan, probably contracted in the USA during his 7-day stay in Los Angeles. As of 27 Sep 2005, California shared 38 percent of the national cumulative cases of West Nile fever in the USA for 2005 (681 of 1804). The index patient recalled having been bitten by "a mosquito" while visiting LA. The odds of getting WNV are normally low, given that only 20

Culex pipiens

C. tritaeniorhynchus

C. vishnui

Aedes albopictus

Ae. dorsalis

Ae. japonicus

Middle East :

the Greek biographer Plutarch reports : "When Alexander the Great arrived before the walls of Babylon he saw a large number of ravens flying about and pecking one another, and some of them fell dead in front of him". The ravens might have been dying of WNV infection as they belong to a family of birds that are particularly susceptible to the pathogen (members of the same family are responsible for WNV spread across the USA) and this could explain his sudden death in 323 BC, when he aged only 32, which has puzzled historians for years (poisoning, flu and typhoid fever have all been suggested, based on records of his 2-week illness of infection, liver disorder, rash)
Israel, the only land bridge between 3 continents, is at a junction for birds migrating south from Eurasia to Africa in autumn and north to their breeding grounds in spring. In spring the Red Sea and the Gulf of Aqaba/Eilat act as a long deflection barrier diverting many northbound migrants to Eilat (Shirihai and Christie 1992). Eilat is strategically located at the northern edge of almost 2000 km of continuous desert regions of the Sinai, Sahara and the Sahel and many of the avian migrants land here to rest and refuel after having crossed the inhospitable deserts to the south (Safriel 1968, Yosef 1998). Since 1984 a trapping and ringing has been conducted by the IBRCE at Eilat. Following the recent interest in migratory birds as vectors of WNV in the region we sampled birds migrating to Africa in autumn 2001 and upon their return to Europe in spring 2002. The species that stood out as having one of the highest levels of antibodies was the Little Stint (Calidris minuta). In spring, of 82 individuals sampled only 2 (2.4%) were positive. In contrast in autumn 2001 we sampled 177 individuals of which 18 (10.2 percent) were positive. This reversal of results is further confounded by the fact that 16 of the 18 Little Stint were 1st year birds, i.e., individuals that had recently fledged and were on their 1st migration south. Hannoun et al (1972) also found that one (20 percent) of the 5 Little Stint they sampled in Tunisia was seropositive. The Little Stint is a monotypic species in which it is easy to separate the juveniles from adults based on plumage. The Little Stint breeds on high-arctic coastal mainland tundra and winters mainly in Africa, around the Indian Ocean and on coasts of the Indian sub-continent, with variable numbers also in the Mediterranean basin and the Persian Gulf (Cramp 1983). In conclusion, based on our data, and the lack of any other studies on the species or other tundra breeding avian species, we suggest that the arctic region has a greater level of viremia and for the transfer of WNV between rganisms than has previously been realized. Future studies not assume blindly that the virus is out of Africa alone. But the report is rife with assumptions: (1) Migrating birds are important intercontinental "vectors" of WNV; (2) Seropositive migrant hatch-year sandpipers (stints) were exposed on their breeding grounds in the Arctic; and (3) Mosquitoes are responsible for local virus spread. All 3 of these assumptions can be debated, and none are well-established scientific facts. For example, Little Stints probably leave their breeding sites in July and may be captured in Eilat in August-September. Thus they migrate several thousand miles over an extended period of time, resting frequently in wetlands, many of which could be WNV transmission foci. It is even possible that the Little Stints captured in Eilat had been present for several weeks in Eilat, where WNV has been reported to be endemic. Before the scientific community starts assuming that WNV is transmitted in the arctic (where no Culex mosquitoes are present), better evidence is needed. The main problem with serological test results from migratory birds, aside from the difficulties and complexities of flavivirus serology, is that all such birds have unknown travel histories, and thus the site of infection is unknown. These hatch-year Little Stints could have long-lasting maternal antibody, though this is unlikely. Arctic mosquitoes are notorious pests and can occur in tremendous numbers. Most are species of Aedes and Ochlerotatus. Culex mosquitoes are generally absent from such northern areas but Culex torrentium , an ornithophagic species, is found north of the arctic circle in Scandanavia. Culex territans, which feeds mainly on reptiles and amphibia, but also possibly on mammals, is found in Alaska and the North West Territories of Canada. And, there is at least one record of Culex tarsalis being found in the North West Territories of Canada near the MacKenzie valley. So a few Culex mosquitoes are found in the far northern areas. It might be worth remembering that West Nile virus has been isolated from a very great number of mosquito species - in about 11 genera. And in Russia the virus has been isolated from both ixodid ticks (Hyalomma marginatum marginatum) and from argasid ticks (Orithodoros capensis). I highly doubt that the arctic could be a major source of bird infection or transmission for West Nile Virus (WNV) in light of the following factors:

although in Alaska we have a total of 4 mosquito species that have been implicated in some way with the possible transmission of WNV (Culex tarsalis, plus 2 Ochlerotatus and one Culiseta species), most apparently tested positive for WNV but have not necessarily been proven to be competent vectors for the disease. Culex tarsalis, as mentioned in previous posting, is predominantly a reptilian and/or amphibian feeder that feeds on mammals rather than birds as a facultative host, and as such should not be a major factor in WNV transmission at high northern latitudes.
all 35 or so of our described mosquito species in Alaska for which the life history is known have single annual breeding capabilities due to the very short summer breeding season. It should be very difficult to amplify the disease in arctic regions through multiple bird-mosquito-bird transmissions because of this
the worst-case scenario would be that overwintering mosquito species in arctic latitudes would pick up WNV 1st thing in the spring from migrating birds and deliver it to other birds, and the amplification would continue through summer mosquito species. 2 of 4 WNV-implicated mosquito species are active during the spring season, while 2 are summer species. The ranges of these species do not overlap very well in most places, especially in the arctic. Nonetheless, given the way the disease has spread westward within North America, we can no longer say that we in Alaska are entirely immune from possible invasion by WNV. It would be interesting to learn whether WNV can overwinter within the salivary glands of mosquitoes in arctic climates. That might complicate things, but even so, I very much doubt whether WNV is -- or will ever be -- significantly propagated in arctic latitudes in North America.

Culex tarsalis

Culex territans

Culex

Ochlerotatus

Oc. impiger

Oc. nigripes

Cx. tarsalis

Oc. churchillensis

Oc. rempeli

Genomics^{ref1,
ref2,
ref3} :

lineage 1 is composed of WNV strains from different geographic regions, and it is subdivided into at least 3 clades

clade A contains strains from Europe, Africa, the Middle East, and America
clade B represents the Australian (Kunjin) strains
clade C contains Indian WNV isolates

lineage 2 contains the B 956 prototype strain and other strains isolated so far exclusively in sub-Saharan Africa and Madagascar.
lineage 3 consists of a virus strain isolated from Culex pipiens mosquitoes at the Czech Republic/Austria border (named Rabensburg virus)
lineage 4 consists of a unique virus isolated in the Caucasus. These 2 viruses, however, may also be considered independent flaviviruses within the JEV complex^ref

Transmission :

a zooanthroponosis ; reservoir : infected Aves spp. and Equus caballus (although a horse infected with WNV can survive, there have been a number of equine deaths following WNV infection. The horses do become very ill in most cases. 2 equine vaccines preventative for the viral disease have been approved in the USA). Gallus gallus are used as "sentinels" to detect when the virus is present in mosquitoes as a warning for humans, horses, and other animals.

vectors : > 43 species of ornithophilic mosquitoes in 10 genera, including Culex spp. (including Culex pipiens pipiens , Culex pipiens quinquefasciatus , and Culex tarsalis ) and Hippoboscid spp., have been found positive for WNV in the USA since 1999. Humans and other mammals don't seem to carry enough of the virus in their blood to pass it along to birds or other animals through mosquitoes, so if you have a mosquito that only bites mammals, there is no transmission to humans. The virus can also infect ornithophagic ticks (Argas hermanni and Hyalomma asiaticum), but their role in the epidemiology of WN remains unclear. Nevertheless it appears that Culex spp., in particular Culex pipiens s.s. x Cules pipiens molestus , are the principal vectors in the USA^ref. Elsewhere, such as in Europe^ref and Africa, Culex spp. mosquitoes are the main vectors, including some species that provide a bridge by biting both birds and humans. Because viremia is low, competent vector mosquitoes probably rarely transmit virus from an infected human host.
despite there is no evidence that a person can get infected by handling live or dead infected birds, it is not inconceivable that WNV could be transmitted via wounds and abrasions.

WNV can be transmitted directly between captive birds in the absence of vectors and ectoparasites

Transmission among alligators has been assumed to be by mosquitoes, but it is possible that there is direct transmission via wounding. Conventionally, mosquitoes become infected when feeding on viremic birds and subsequently transmit the virus to susceptible hosts. Nonviremic transmission of WNV between cofeeding mosquitoes has been demonstrated. Donor, Culex pipiens quinquefasciatus mosquitoes infected with WNV were fed simultaneously with uninfected "recipient" mosquitoes on naïve mice. At all times, donor and recipient mosquitoes were housed in separate sealed containers, precluding the possibility of mixing. Recipients became infected in all 5 trials, with infection rates as high as 5.8% and no detectable viremia in the hosts. Remarkably, a 2.3% infection rate was observed when 87 uninfected mosquitoes fed adjacent to a single infected mosquito. This phenomenon could potentially enhance virus survival, transmission, and dispersion and obviate the requirement for viremia. All vertebrates, including immune and insusceptible animals, might therefore facilitate mosquito infection. These findings question the status of dead-end hosts in the WNV transmission cycle and may partly explain the success with which WNV established and rapidly dispersed throughout North America

^ref

WNV is not transmitted from person to person due to low viremia, excepted :

transplacental transmission
colostral transmission (CDC has reported a single case of mother-to-infant infection and a second transmission to a breastfed child)
blood transfusion-associated transmission (TAT) (tested by Procleix^®) : in June 2003, blood-collection agencies (BCAs) implemented investigational WNV nucleic acid-amplification tests (NATs) performed on mini-pools (MP NAT) of samples from 6 or 16 donations^ref (when positive or if in areas with epidemic activity => individual donation testing (IDT NAT) to screen all blood donations and identify potentially infectious donations for quarantine and retrieval. A donation that was repeatedly reactive on IDT NAT was considered to be from a presumptive viremic donor (PVD). As of 31 Mar 2004, state and local health departments had reported 818 PVDs to ArboNET; dates of collection ranged from 25 Jun to 2 Dec 2003. Complete information was available for 811 (99%) of these PVDs; 6 (1%) had West Nile viral encephalitis or meningitis subsequent to donation (median age: 45 years, range: 28 to 76), 137 (17%) had West Nile fever (median age: 46 years, range: 17 to 76), and 654 (81%) remained asymptomatic. Of the PVDs reported to ArboNET, 691 (85%) were residents of 9 states (Colorado, Kansas, Nebraska, New Mexico, North Dakota, Oklahoma, South Dakota, Texas, and Wyoming). These states experienced WNV epidemics in 2003 and accounted for 60% of reported cases of West Nile viral encephalitis or meningitis. TAT of West Nile virus (WNV) in the United States was 1st identified in 2002^ref. During 2003, a total of 6 confirmed or probable cases of WNV TAT were reported to CDC. The majority of BCAs use a 2-tiered NAT-screening algorithm. On the basis of the test manufacturer's format, NATs are conducted on minipools of samples from either 6 or 16 blood donations. If a minipool is nonreactive, its constituent donations are released for transfusion. If a minipool is reactive, the constituent donations undergo individual testing. If an individual donation is reactive, associated blood components are impounded, and the donor is notified for further testing to confirm the infection. In 2003, blood-donation screening for WNV resulted in the impounding of approximately 800 blood components potentially containing WNV. However, 6 reported cases of transfusion-associated WNV disease were associated with units of blood components with viral concentrations too small to be detected by minipool NAT^ref. In 2004, to improve the sensitivity of WNV screening, BCAs implemented systems to trigger a switch from minipool NAT to individual NAT in areas with epidemic WNV transmission. This report describes the 1st transfusion-associated WNV infection identified in 2004; the implicated blood donation was collected before the switch to individual testing. Clinicians should remain aware of the risk for WNV transmission through blood-product transfusion and alert state health officials to hospitalized patients with WNV disease symptoms who have had a transfusion during the preceding 28 days. On admission, he was febrile, had altered mental status, oscillopsia, and cogwheel rigidity. Magnetic resonance imaging of the brain was consistent with WNV encephalitis^ref. For this reason, BCAs developed systems to trigger a switch from minipool to individual NAT in areas of epidemic WNV transmission^ref. Evidence of year-round WNV activity has been documented in east Texas and Louisiana^ref. Routine testing in 2003 and 2004 identified 540 donations that were positive for WNV RNA, of which 362 (67%) were IgM-antibody–negative and most likely infectious. Of the 540 positive donations, 148 (27%) were detectable only by testing of individual donations, but only 15 of the 148 (10%) were negative for IgM antibody. The overall frequencies of RNA-positive donations during the epidemic periods were 1.49 per 10,000 donations in 2003 and 0.44 per 10,000 in 2004. In 2004, 52 percent of the positive donations were from donors in four counties in southern California. Rapid implementation of a NAT led to the prospective identification of 519 donors who were positive for WNV RNA and the removal of > 1000 potentially infectious related components from the blood supply of the Red Cross. No cases of transfusion-transmitted infection were confirmed among recipients of the tested blood^ref. Although nucleic acid amplification testing of minipools of blood donations prevented hundreds of cases of West Nile virus infection in 2003, it failed to detect units with a low level of viremia, some of which were antibody-negative and infectious. These data support the use of targeted nucleic acid amplification testing of individual donations in high-prevalence regions, a strategy that was implemented successfully in 2004^ref.
tissue transplantation : in September 2005, WNV infection was confirmed in 3 of 4 recipients of organs transplanted from a common donor. 2 recipients subsequently had neuroinvasive disease, one recipient had asymptomatic WNV infection, and a 4th recipient apparently was not

infected. The organ donor, a New York City resident, was hospitalized on 23 Aug 2005 after a traumatic head injury and underwent emergency evacuation of an epidural hematoma, during which he received one unit of packed red blood cells (PRBCs). He was declared brain dead on 26 Aug 2005. Liver and associated vessels, one lung, and both kidneys were recovered. On 28 Aug 2005, the liver and kidneys were transplanted into 3 recipients at 2 transplant centers in New York City, the lung was transplanted into a recipient at a transplant center in Pittsburgh, and the vessels were discarded. After unexplained neurologic illness occurred in 2 organ recipients, an investigation was initiated. Investigators determined that the donor had lived near an area where mosquitoes positive for WNV were collected on 16 Aug 2005. The donor's wife reported that he had spent time outdoors and felt febrile before sustaining the fatal head injury. Serum and plasma collected from the donor on 27 Aug 2005 were retrieved. The samples tested positive for WNV immunoglobulin M antibodies (IgM) and IgG by enzyme immunoassay but negative for WNV RNA by polymerase chain reaction (RT-PCR). Immunohistochemical analyses of liver, gallbladder, kidney, and epidural hematoma were negative for WNV antigens. The PRBC unit received by the organ donor was donated on 30 Jul 2005 and was negative for WNV RNA by minipool nucleic acid-amplification test (mpNAT). A repeat donation on 22 Sep 2005 was WNV mpNAT and IgM negative.

the liver recipient had end-stage liver disease caused by hepatitis C virus infection. She initially did well after the transplantation. She required multiple transfusions of blood products, all of which were WNV RNA negative by mpNAT. On post-transplant day 13, she had a fever and altered mental status. On day 18, she experienced respiratory distress requiring endotracheal intubation. A lumbar puncture revealed mild lymphocytic pleocytosis (8 cells/mm3) and elevated protein (81 mg/dL). She became comatose and developed acute flaccid paralysis consistent with WNV encephalitis. Serum and cerebrospinal fluid (CSF) specimens collected on day 23 were positive for WNV IgM, and CSF contained WNV RNA. That day, the patient began treatment with 4 doses of intravenous Omr-IgG-amTM (Omrix Biopharmaceuticals, Tel Aviv, Israel, supplied by the National Institutes of Health [NIH]), an immune globulin with high antibody titers against WNV under an investigational new drug (IND) compassionate-use protocol; however, the patient had no subsequent clinical improvement and remains in a coma.
the lung recipient had end-stage lung disease caused by pulmonary fibrosis. The initial post-transplant course was uneventful aside from blood-product receipt. The patient went home on post-transplant day 16 but was re-admitted the following day with fever and dyspnea requiring endotracheal intubation, followed by altered mental status, seizures, and acute flaccid paralysis consistent with WNV encephalitis. On day 23, a lumbar puncture revealed elevated CSF protein (149 mg/dL) but no white blood cells; a brain magnetic resonance image taken the same day was normal. Serum collected on day 19 was negative for WNV IgM, but, by day 23, serum was IgM and IgG positive. CSF from day 24 was negative for WNV IgM and WNV RNA, but CSF from day 27 was positive for WNV IgM and IgG. The patient completed experimental treatment with 4 doses of Omr-IgG-am, without clinical improvement, and remains in a coma.
the 1st kidney recipient had end-stage renal disease attributable to IgA nephropathy. She had no immediate post-transplant complications, received no blood products, and was discharged home on day 3. Serum collected on day 22 was negative for WNV IgM but positive for IgG (consistent with a previous flavivirus infection) and was positive for WNV RNA. The patient was re-admitted to the hospital on day 27 for experimental Omr-IgG-am treatment and remains asymptomatic.
the 2nd kidney recipient had end-stage renal disease caused by Alport syndrome. He received blood products after the transplant and was discharged home on post-transplant day 7. Serum collected from the patient on day 16 was negative for WNV IgM, IgG, and RNA. As a precaution, the patient was re-hospitalized on day 27 for experimental Omr-IgG-am treatment. He remains well.

This report describes the 2nd case of WNV transmission associated with organ transplant

^ref

. Several important differences exist between this and the previously reported occurrence. The 1st organ donor-associated WNV transmission, reported in August 2002, occurred after the donor received a transfusion of WNV⁺ blood one day before organ recovery. A serum sample collected immediately before organ recovery subsequently tested positive for WNV by PCR and culture but lacked WNV IgM antibodies. All 4 organ recipients were infected and became ill. In contrast, the current organ donor was likely infected via a mosquito bite rather than through blood transfusion, and a serum sample obtained one day before the organs were recovered had WNV IgM and IgG antibodies but was PCR negative. The lung and liver transplant recipients had severe WNV encephalitis and acute flaccid paralysis with respiratory failure; one kidney recipient had a positive PCR test result in serum 22 days after transplantation and remains asymptomatic, and the other kidney recipient had no evidence of WNV infection. Serologic and clinical studies indicate that organ-transplant recipients have a risk approximately 40 times that of the general population for neuroinvasive disease after WNV infection

^ref

. Infected organ-transplant recipients and other immunosuppressed persons typically have prolonged WNV incubation periods, as judged from a longer time than normal needed for the emergence of specific immunoglobulins, during which asymptomatic viremia can be detected

^ref

. The infected kidney recipient had symptomatic viremia 22 days after transplant. All of the recipients were treated through a Food and Drug Administration (FDA)-approved IND compassionate-use protocol with Omr-IgG-am, an intravenous immunoglobulin product with high-titered neutralizing antibody to WNV. No proven effective treatment or prophylaxis for WNV infection exists; a randomized placebo-controlled, double-blind trial of Omr-IgG-am is under way

^ref

. Investigation of 30 recognized cases of WNV transmitted by blood transfusion documented to date indicated that the donors' viremias can be of low titer and that all resulted from IgM antibody-negative donations

^ref

. Conversely, transfused viremic donations that were recognized only after retrospective testing did not transmit WNV infection if IgM antibody was present

^ref

. Since 2003, the U.S. blood supply has been screened for WNV using NAT, which has reduced the risk for transfusion transmission

^ref

. The organ-transplant-associated WNV transmission described in this report suggests that transmission through solid organ transplantation can occur from donors with IgM and IgG antibodies and without detectable nucleic acid by PCR in their serum. Experimental evidence in humans and animals suggests that WNV might persist in organs after clearance of viremia

^ref

. Further testing of the donor serum using a highly sensitive NAT assay for blood-donor screening is pending. Organ donors are screened to identify infectious risks on the basis of national organ-procurement standards

^ref

. Screening of all organ donors with WNV NAT is not currently required or routinely performed because of 1) NAT availability only through IND applications for blood screening, 2) the length of turnaround time to obtain WNV NAT testing, and 3) the unproven test performance on donated organs. One analysis suggested that WNV NAT screening might result in a net loss of years of life among certain types of potential transplant recipients

^ref

by excluding healthy donors from an already limited donor pool. National guidelines for organ-donor screening are continuously reevaluated by the Health Resources and Services Administration in consultation with FDA, CDC, and organ-procurement organizations

^ref

Risk factors : who practiced >2 personal protective behavior traits (avoidance of exposure to mosquitoes, wearing long sleeves and pants, using mosquito repellent) had >= 50% reduction in risk of infection. Time spent outside at dusk or dawn on a nonwork day was a significant risk factor for WNV infection, which is consistent with findings from a previous report^ref. Finding mosquitoes in the home was not associated with WNV infection, as it was in a previous report^ref1,
ref2.
=> West Nile fever (WNF) :

West Nile asymptomatic infection (WNAI) (80%) but are believed to have viremia lasting a median of 6.5 days^ref1,
ref2. Asymptomatic WNV-infected people with viremia probably represent the largest risk group of blood donors. Because symptom screening at the time of blood donation will not identify most viremic donors, screening by NAT was implemented rapidly to identify potentially infectious blood donations by detecting WNV RNA
symptomatic (20%) : fever , drowsiness, severe frontal headache , eye floaters, maculopapular exanthema ^ref, abdominal pain, loss of appetite, nausea, and generalized lymphadenopathy

self-limiting within 3-5 days (80%)
neuroinvasion (< 1%^ref) evolving to sometimes lethal West Nile neurological syndromes (WNNS) : acute West Nile meningoencephalitis , WN meningitis , WN encephalitis , and WN myelitis / polio-like syndrome (20% : probably in patients with hypofunctional isoforms of 2',5'-oligoadenylate synthetase )

Laboratory examinations : both a positive antibody result with ELISA and hemagglutination tests can be induced by other flaviviruses and certain immunizations (for example, yellow fever, tickborne encephalitis, Japanese encephalitis, and St Louis encephalitis), and so it is necessary to determine whether an immunization or infection with one of these agents could be the cause. When suspected, a neutralization test is still necessary.
Therapy : neutralization of WNV in vivo correlates with the development of an antibody response against the viral envelope (E) protein. Using random mutagenesis and yeast surface display, individual contact residues of 14 newly generated monoclonal antibodies against domain III of the WNV E protein have been defined. MAbs that strongly neutralized WNV localized to a surface patch on the lateral face of domain III. Convalescent antibodies from individuals who had recovered from WNV infection also detected this epitope. E16 mAb neutralized 10 different strains in vitro, and showed therapeutic efficacy in mice, even when administered as a single dose 5 d after infection. A humanized version of E16 was generated that retained antigen specificity, avidity and neutralizing activity. In postexposure therapeutic trials in mice, a single dose of humanized E16 protected mice against WNV-induced mortality, and may therefore be a viable treatment option against WNV infection in humans^ref.
Prognosis : mortality < 0.1%
Prevention : currently the only vaccine against WNV is an inactivated one manufactured by Fort Dodge and approved for use in equines in 2 doses
Web resources :

WNV, CDC
National and State West Nile Virus Maps - Human - USA by US Geological Survey, Center for Integration of Natural Disaster Information (CINDI)
West Nile Virus Maps and Statistics by National Pesticide Information Center (NPIC)