wildlife and zoo assay data sheet
West Nile virus (WNV)
Ultrasensitive qualitative detection of West Nile virus by
reverse transcription coupled real time polymerase chain reaction
virus (WNV) belongs to the genus Flavivirus of the family
Flaviviridae and is an arthropod-borne virus. It possesses a
single-stranded plus-sense RNA genome of approximately 11,000
nucleotides. It circulates in natural transmission cycles
involving primarily Culex species mosquitoes and birds; humans
and other mammals, such as primates, are thought to be
Historically, WNV was found primarily in Africa, Asia, southern
Europe, and Australia and was responsible for several
significant epidemics, notably, in Israel (1950s), France
(1962), South Africa (1974), and Romania (1996) (Hayes, 1989;
Tsai et al., 1998;Savage et al., 1999). In 1999 and 2000, WNV
was responsible for epidemics and epizootics in the northeastern
United States, in which there were human fatalities and
extensive avian mortality (Anderson et al., 1999; Lanciotta et
al., 1999). On the basis of retrospective serosurveys conducted
in New York City in 1999 and 2000, symptomatic illness develops
in approximately 20% of persons infected with WNV and
approximately 1 in 150 human infections results in
meningoencephalitis, the most commonly reported form of
In 2002, an
outbreak of West Nile virus infection occurred in the state of
Louisiana in which 319 human cases of WNV-associated illness
were reported. Most of these cases happened in the southeastern
portion of the state, including St. Tammany Parish. The Tulane
National Primate Research Center (TNPRC) is located in St.
Tammany Parish and houses large outdoor breeding colonies of
baboons and macaques. A serological survey of primates in these
breeding colonies indicated that approximately 36% of the
nonhuman primates were infected with WNV during the 2002
transmission season (Ratterree et al., 2003). Implications of
this study are that nonhuman primates can be as susceptible to
West Nile virus infection as humans, and captive primate
populations can be a potential source of viral carriers.
Surveillance for West Nile virus relies on the testing of
field-collected mosquitoes and on the testing of dead birds for
the presence of virus by isolation in cell culture. However,
virus isolation followed by identification through
immunofluorescence assays can take over a week to complete. In
addition, virus isolation in cell culture from CSF or serum has
generally been unsuccessful, likely due to the low level and
short-lived viremia associated with infections with these
viruses (Monath and Heinz, 1996; Southam and Moore, 1954).
infections can be inferred by immunoglobulin M (IgM) capture and
IgG enzyme-linked immunosorbent assays (ELISAs); however,
confirmation of the type of infecting virus is possible only by
detection of a fourfold or greater rise in virus-specific
neutralizing antibody titers in either cerebrospinal fluid (CSF)
or serum by performing the plaque reduction neutralization assay
(PRNT) with several flaviviruses (Johnson et al., 2000; Martin
et al., 2000). Thus serological detection of WNV infection is
neither specific nor sensitive. PCR
detection of West Nile virus is now considered to be a
rapid, specific and sensitive detection method to identify this
Help confirm the disease causing agent
Help ensure that animal groups and populations are free of
West Nile Virus
Early prevention of spread of the virus among a
Minimize human exposure to the virus
Anderson, J. F., Andreadis, T.G., Vossbrinck, C.R., Tirrell,
S.,Wakem, E.M., French, R.A., Garmendia, A.E. and Van Kruiningen,
H.J. (1999) Isolation of West Nile virus from mosquitoes, crows,
and a Cooper's hawk in Connecticut. Science 286:2331-2333.
Hayes, C. G. (1989). West Nile fever, p. 59-88. In T. P. Monath
(ed.), The arboviruses: epidemiology and ecology, vol. V. CRC
Press, Inc., Boca Raton, Fla.
Johnson, A. J., Martin, D.A.,
Karabatsos, N. and Roehrig, J.T.(2000) Detection of
antiarboviral immunoglobulin G by using a monoclonal
antibody-based capture enzyme-linked immunosorbent assay. J.
Clin. Microbiol. 38:1827-1831.
Lanciotti, R. S., Roehrig,
J.T., Deubel, V., Smith, J., Parker, M., Steele, K., Volpe,
K.E., Crabtree, M.B., Scherret, J.H., Hall, R.A., MacKenzie,
J.S., Cropp, C.B., Panigrahy, B., Ostlund, E., Schmitt, B.,
Malkinson, M., Banet, C., Weissman, J., Komar, N., Savage, H.M.,
Stone, W., McNamara, T. and Gubler, D.J.(1999) Origin of the
West Nile virus responsible for an outbreak of encephalitis in
the northeastern U.S. Science 286:2333-2337.
Martin, D. A.,
Muth, D.A., Brown, T., Johnson, A.J., Karabatsos, N. and Roehrig,
J.T. (2000) Standardization of immunoglobulin M capture
enzyme-linked immunosorbent assays for routine diagnosis of
arboviral infections. J. Clin. Microbiol. 38:1823-1826.
Monath, T. P., and Heinz, F.X. (1996) Flaviviruses, p. 978-984.
In B. N. Fields (ed.), Fields virology, 3rd ed., vol. 1.
Lippincott-Raven Publishers, Philadelphia, Pa.
M.S., da Rosa, A.P., Bohm, R.P. Jr, Cogswell, F.B., Phillippi,
K.M., Caillouet, K., Schwanberger, S., Shope, R.E. and Tesh,
R.B.(2003) West Nile virus infection in nonhuman primate
breeding colony, concurrent with human epidemic, southern
Louisiana. Emerg Infect Dis. 9:1388-1394.
Southam, C. M.,
and Moore, A.E. (1954) Induced virus infections in man by the
Egypt isolates of West Nile virus. Am. J. Trop. Med. Hyg.
Savage, H. M., Ceianu, C., Nicolescu, G.,
Karabatsos, N.,Lanciotti, R., Vladimirescu, A., Laiv, L.,
Ungureanu, A., Romanca, C. and Tsai, T.F. (1999). Entomologic
and avian investigations of an epidemic of West Nile fever in
Romania, 1996, with serological and molecular characterization
of a virus from mosquitoes. Am. J. Trop. Med. Hyg. 61:600-611.
Tsai, T. F., Popovici, F., Cernescu, C., Campbell, G.L. and
Nedelcu, N.I. (1998) West Nile encephalitis epidemic in
southeastern Romania. Lancet 352:767-771.
- 0.2 ml CSF, or 0.2 ml fresh or frozen CNS tissue.
- 0.2 ml whole blood in EDTA (purple top) or ACD (yellow top)
tube, or 0.2 ml serum or plasma.
Contact Zoologix if advice is needed to determine an appropriate specimen type for a specific diagnostic application. For specimen types not listed here, please contact Zoologix to confirm specimen acceptability and shipping instructions.
specimen types, if there will be a delay in shipping, or during
very warm weather, refrigerate specimens until shipped and ship
with a cold pack unless more stringent shipping requirements are
specified. Frozen specimens should be shipped so as to remain
frozen in transit. See shipping
instructions for more information.
2 business days
Qualitative reverse transcription coupled real time PCR