West nile virus laboratory tests: Difference between revisions
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{{West nile virus}} | {{West nile virus}} | ||
{{CMG}} | {{CMG}}; {{AE}} {{Rim}} | ||
==Overview== | ==Overview== | ||
The front-line screening assay for laboratory diagnosis of human West Nile virus (WNV) infection is the [[IgM]] assay. IgM and IgG ELISA tests can cross-react between [[flavivirus]]es; therefore, serum samples that are antibody-positive on initial screening should be evaluated by a more specific test; currently the plaque reduction neutralization test (PRNT) is the recommended test for differentiating between flavivirus infections. Specimens submitted for WNV testing should also be tested by ELISA and PRNT against other arboviruses known to be active or be present in the area or in the region where the patient traveled. | The front-line screening assay for laboratory diagnosis of human West Nile virus (WNV) infection is the [[IgM]] assay. IgM and IgG ELISA tests can cross-react between [[flavivirus]]es; therefore, serum samples that are antibody-positive on initial screening should be evaluated by a more specific test; currently the plaque reduction neutralization test (PRNT) is the recommended test for differentiating between flavivirus infections. Specimens submitted for WNV testing should also be tested by ELISA and PRNT against other arboviruses known to be active or be present in the area or in the region where the patient traveled. Numerous procedures have been developed for detecting viable WNV, WNV antigen or WNV RNA in human diagnostic samples. These procedures vary in their [[sensitivity]], [[specificity]], and time required to conduct the test. Among the most sensitive procedures for detecting WNV in samples are those using RT-[[PCR]] to detect WNV RNA in human CSF, serum and other tissues. Confirmation of virus isolate identity can be accomplished by indirect [[immunofluorescence assay]] (IFA) using virus-specific monoclonal antibodies or nucleic acid detection. [[Immunohistochemistry]] (IHC) using virus-specific MAbs on brain tissue has been very useful in identifying cases of WNV infection.<ref name=CDC><West Nile Virus in the United States: Guidelines for Surveillance, Prevention, and Control. 4th revision, 2013. [http://www.cdc.gov/westnile/resources/pdfs/wnvGuidelines.pdf] </ref> | ||
==Laboratory Tests== | ==Laboratory Tests== | ||
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Minimally, onset and sample collection dates are required to perform and interpret initial screening tests. The remaining information is required to evaluate any specimens positive on initial screening. If possible, a convalescent [[serum]] sample taken at least 14 days following the acute sample should be obtained to enable confirmation by serological testing.<ref name=CDC><West Nile Virus in the United States: Guidelines for Surveillance, Prevention, and Control. 4th revision, 2013. [http://www.cdc.gov/westnile/resources/pdfs/wnvGuidelines.pdf] </ref> | Minimally, onset and sample collection dates are required to perform and interpret initial screening tests. The remaining information is required to evaluate any specimens positive on initial screening. If possible, a convalescent [[serum]] sample taken at least 14 days following the acute sample should be obtained to enable confirmation by serological testing.<ref name=CDC><West Nile Virus in the United States: Guidelines for Surveillance, Prevention, and Control. 4th revision, 2013. [http://www.cdc.gov/westnile/resources/pdfs/wnvGuidelines.pdf] </ref> | ||
===Serology=== | ===Serology=== | ||
====IgM and IgG ELISA Tests==== | ====IgM and IgG ELISA Tests==== | ||
Revision as of 01:45, 12 September 2014
Editor-In-Chief: C. Michael Gibson, M.S., M.D. [2]; Associate Editor(s)-in-Chief: Rim Halaby, M.D. [3]
Overview
The front-line screening assay for laboratory diagnosis of human West Nile virus (WNV) infection is the IgM assay. IgM and IgG ELISA tests can cross-react between flaviviruses; therefore, serum samples that are antibody-positive on initial screening should be evaluated by a more specific test; currently the plaque reduction neutralization test (PRNT) is the recommended test for differentiating between flavivirus infections. Specimens submitted for WNV testing should also be tested by ELISA and PRNT against other arboviruses known to be active or be present in the area or in the region where the patient traveled. Numerous procedures have been developed for detecting viable WNV, WNV antigen or WNV RNA in human diagnostic samples. These procedures vary in their sensitivity, specificity, and time required to conduct the test. Among the most sensitive procedures for detecting WNV in samples are those using RT-PCR to detect WNV RNA in human CSF, serum and other tissues. Confirmation of virus isolate identity can be accomplished by indirect immunofluorescence assay (IFA) using virus-specific monoclonal antibodies or nucleic acid detection. Immunohistochemistry (IHC) using virus-specific MAbs on brain tissue has been very useful in identifying cases of WNV infection.[1]
Laboratory Tests
Appropriate selection of diagnostic procedures and accurate interpretation of findings requires information describing the patient and the diagnostic specimen. For human specimens, the following data must accompany sera, CSF or tissue specimens for results to be properly interpreted and reported:[1] 1) Symptom onset date (when known) 2) Date of sample collection 3) Unusual immunological status of patient (e.g., immunosuppression) 4) State and county of residence 5) Travel history (especially in flavivirus-endemic areas) 6) History of prior vaccination (e.g., yellow fever, Japanese encephalitis, or Tick-borne encephalitis viruses) 7) Brief clinical summary including clinical diagnosis (e.g., encephalitis, aseptic meningitis).
Minimally, onset and sample collection dates are required to perform and interpret initial screening tests. The remaining information is required to evaluate any specimens positive on initial screening. If possible, a convalescent serum sample taken at least 14 days following the acute sample should be obtained to enable confirmation by serological testing.[1]
Serology
IgM and IgG ELISA Tests
The front-line screening assay for laboratory diagnosis of human WNV infection is the IgM assay. Currently, the FDA has cleared four commercially-available test kits from different manufacturers, for detection of WNV IgM antibodies. These four kits are used in many commercial and public health laboratories in the United States. In addition the CDC-defined IgM and IgG ELISA can be used; protocols and reagents are available from the CDC DVBD Diagnostic Laboratory. There is also a microsphere-based immunoassay for the detection of IgM antibodies that can differentiate WNV from SLE.[1]
Plaque Reduction Neutralization Test
Because the IgM and IgG ELISA tests can cross-react between flaviviruses (e.g., SLE, dengue, yellow fever, WN), they should be viewed as screening tests only. For a case to be considered confirmed, serum samples that are antibody-positive on initial screening should be evaluated by a more specific test; currently the plaque reduction neutralization test (PRNT) is the recommended test for differentiating between flavivirus infections. Though WNV is the most common cause of arboviral encephalitis in the United States, there are several other arboviral encephalitides present in the country and in other regions of the world. Specimens submitted for WNV testing should also be tested by ELISA and PRNT against other arboviruses known to be active or be present in the area or in the region where the patient traveled.[1]
Virus Detection Assay
Numerous procedures have been developed for detecting viable WNV, WNV antigen or WNV RNA in human diagnostic samples. These procedures vary in their sensitivity, specificity, and time required to conduct the test.[1]
RT-PCR
Among the most sensitive procedures for detecting WNV in samples are those using RT-PCR to detect WNV RNA in human CSF, serum and other tissues. Fluorogenic 5' nuclease techniques (real-time PCR) and nucleic acid sequence-based amplification (NASBA) methods have been developed and validated for specific human diagnostic applications and for detecting WNV RNA in blood donations.[1]
WNV presence can be demonstrated by isolation of viable virus from samples taken from clinically ill humans. Appropriate samples include CSF (serum samples may be useful very early in infection) and brain tissue (taken at biopsy or postmortem). Virus isolation should be performed in known susceptible mammalian (e.g., Vero) or mosquito cell lines (e.g., C6/36). Mosquito origin cells may not show obvious cytopathic effect and must be screened by immunofluorescence or RT-PCR. Appropriate samples for virus isolation from clinically ill humans include CSF (serum samples may be useful very early in infection) and brain tissue (taken at biopsy or postmortem).[1]
Indirect Immunofluorescence Assay
Confirmation of virus isolate identity can be accomplished by indirect immunofluorescence assay (IFA) using virus-specific monoclonal antibodies or nucleic acid detection. The IFA using well-defined murine monoclonal antibodies (MAbs) is an efficient, economical, and rapid method to identify flaviviruses. MAbs are available that can differentiate WNV and SLE virus from each other and from other flaviviruses. Incorporating MAbs specific for other arboviruses known to circulate in various regions will increase the rapid diagnostic capacities of state and local laboratories. Nucleic acid detection methods including RT-PCR, TaqMan and nucleic acid sequence based amplification (NASBA) methods may be used to confirm virus isolates as WNV.[1]
While these tests can be quite sensitive, virus isolation and RT-PCR to detect WNV RNA in sera or CSF of clinically ill patients have limited utility in diagnosing human WNV neuroinvasive disease due to the low level viremia present in most cases at the time of clinical presentation. However, one study demonstrated that combining detection of IgM with detection of WNV RNA in plasma significantly increased the number of WNV non-neuroinvasive (i.e., fever) cases detected. Virus isolation or RT-PCR on serum may be helpful in confirming human WNV infection in immunocompromised patients when antibody development is delayed or absent.[1]
Immunohistochemistry
Immunohistochemistry (IHC) using virus-specific MAbs on brain tissue has been very useful in identifying both human and avian cases of WNV infection. In suspected fatal cases, IHC should be performed on formalin fixed autopsy, biopsy, and necropsy material, ideally collected from multiple anatomic regions of the brain, including the brainstem, midbrain, and cortex.[1]