Phylogenetic Relationships of Southern African West Nile Virus Isolates

Felicity J. Burt, Antoinette A. Grobbelaar, Patricia A. Leman, Fiona S. Anthony, Georgina V.F. Gibson, and Robert Swanepoel


Emerging Infectious Diseases. 2002;8(8) 

In This Article

Abstract and Introduction

Phylogenetic relationships were examined for 29 southern African West Nile virus (formal name West Nile virus [WNV]) isolates from various sources in four countries from 1958 to 2001. In addition sequence data were retrieved from GenBank for another 23 WNV isolates and Kunjin and Japanese encephalitis viruses. All isolates belonged to two lineages. Lineage 1 isolates were from central and North Africa, Europe, Israel, and North America; lineage 2 isolates were from central and southern Africa and Madagascar. No strict correlation existed between grouping and source of virus isolate, pathogenicity, geographic distribution, or year of isolation. Some southern African isolates have been associated with encephalitis in a human, a horse, and a dog and with fatal hepatitis in a human and death of an ostrich chick.

West Nile virus (formal name West Nile virus [WNV]) is a mosquito-borne member of the Flaviviridae family (genus Flavivirus), which was originally isolated from the blood of a febrile patient in Uganda in 1937.[1] The virus is widely distributed in Africa, Asia, and Europe and was recently spread to the Western Hemisphere, where its presence was recognized in the northeastern United States in 1999.[2,3]

After the initial isolation of the virus, sporadic cases and outbreaks of febrile disease were recorded in humans in Africa, the Near East, and Asia; the largest outbreaks occurred in Israel in 1950-1954 and 1957 and in South Africa in 1974.[4,5] Meningoencephalitis was first observed in elderly patients in Israel in 1957 and subsequently observed as a complication in young children in India.[6,7] In 1962, the isolation of WNV from a horse with encephalitis was reported in Egypt; from 1962 through 1966, meningoencephalitis occurred in both humans and horses in a series of outbreaks in southern France.[8,9] In 1983, four cases of hepatitis, two fatal, were attributed to WNV infection in the Central African Republic (CAR), a report that has been largely overlooked.[10] A marked increase in the frequency and severity of outbreaks of human disease during the 1990s followed, often involving horses; epidemics occurred in Algeria, Romania, Morocco, Tunisia, Italy, Russia, France, Israel, and the United States.[11,12,13,14,15,16,17,18] Moreover, the recent outbreaks in Romania, Israel, and the United States were characterized by concurrent deaths in birds.[19,20] The virus circulating in the United States was found to be most closely related genetically to a WNV isolate associated with goose deaths in Israel in 1998, suggesting that the virus was imported into America from the Near East, either in an infected bird, mosquito, human, or other animal. The exact mechanism of the introduction will probably remain unknown.[18,19]

In southern Africa, WNV was found to be widely endemic in areas where the principal vector, Culex univittatus, and avian hosts of the virus are present. Human infections tended to be sporadic; large epidemics occurred when unusually high rainfall or hot weather favored breeding of the vectors.[21,22,23] Outbreaks were associated with concurrent epizootics in birds, as evidenced by antibody studies, and 13 species of experimentally infected wild birds supported replication of the virus without becoming overtly sick or dying.[24,25,26] The largest epidemic occurred in 1974 and involved tens of thousands of human cases over a 2,500-km2 area of the Karoo and Northern Cape Provinces.[5,21] A mean antibody prevalence of 55% in humans was recorded in the affected region after the outbreak; levels of 80% to 85% were recorded in some locations. In one town, 1,700 people sought medical attention. Infections were most frequently subclinical or associated with mild febrile illness characterized by rash, myalgia, and arthralgia. No human deaths were recorded, and no excess bird deaths were observed, although an antibody prevalence of 53% was detected in wild birds after the outbreak.[5,21] A smaller epidemic occurred from 1983 through 1984 in association with an outbreak of Sindbis virus (formal name: Sindbis virus) infection in the Witwatersrand-Pretoria region of South Africa, and again no deaths were recorded.[23] Since then, the number of human WNV infections confirmed in South Africa, mainly on the basis of detection of antibody response, has remained fairly constant at approximately 5-15 cases per year. Only a proportion of suspected cases are subject to laboratory investigation. Despite the apparently low level of virus activity, however in recent years, a few isolations of WNV have been made from patients with severe disease, including a fatal case of hepatitis in 1989 and nonfatal encephalitis in 2001 (National Institute for Communicable Diseases [NICD], unpub. data).

The apparent increases in the frequency of neurologic infections, human case-fatality rates, and horse and bird deaths in the Northern Hemisphere raised the question of whether a recent emergence of WNV strains with increased pathogenicities occurred or whether the virulence of the virus had previously been underestimated. Investigations with hemagglutination-inhibition kinetics, titer ratios from cross-neutralization tests, reactivity to monoclonal antibodies, and cDNA/RNA heteroduplex restriction enzyme digest profiles confirmed that strain differences occur but did not establish any links between variants and pathogenicity.[27,28,29,30] More recently, phylogenetic analyses based on different regions of the genome have shown that WNV isolates form two well-supported lineages.[19,31,32] Lineage 1 includes viruses from Africa north of the equator, Europe, Asia, and North America; Kunjin virus (formal name: Kunjin virus [KUNV]) from Australia constitutes a subtype of this lineage. Lineage 2 consists solely of viruses from Africa and Madagascar. These findings support the emergence of increased virulence in lineage 1. Lineage 2 isolates are thought to be associated with endemic infection of low virulence in Africa.[18,19]

The South African prototype WNV isolate, H 442, was obtained in 1958 from the blood of a person with mild febrile disease who had been bitten by mosquitoes while catching birds in mist nets for arbovirus studies.[33] This isolate is the only one from South Africa to have been included in a phylogenetic study, and its characterization as a member of lineage 1[19] seems to be inconsistent with findings for other African isolates from south of the equator. However, apart from recent isolation of WNV from severe cases of human disease in southern Africa, isolations of the virus were associated with a fatal infection in a dog, a horse, and an ostrich chick;[34] unpub. data, NICD). Hence, we were prompted to undertake phylogenetic investigation of southern African WNV isolates.


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