Lab Diagnosis of Yellow fever
المؤلف:
Baijayantimala Mishra
المصدر:
Textbook of Medical Virology
الجزء والصفحة:
2nd Edition , p295
2025-12-21
26
Laboratory confirmation is required in all suspected YF cases. The detection of YF infection can be made from blood and tissue samples by isolation of virus, viral RNA detection and antigen detection and serology from blood.
Yellow fever virus being a level 3 pathogen, the specimens should be handled in biosafety level-3 (BSL-3). As an additional precaution, it is recommended to handle the samples from patients with hemorrhagic manifestations in BSL-4 lab. Laboratory staff should be prior vaccinated for YF. Because of these limitations, the laboratory testing is performed in national or regional reference laboratory and as the disease is more common in rural areas, laboratory confirmation facility is limited in both epidemic and endemic settings.
Isolation of virus: Like many other arboviruses, YFV isolation can be done in cell lines, mosquito inoculation and suckling mice intracerebral inoculation. Isolation of virus can be done from blood or infected tissue samples. Blood should be collected within first few days of illness.
Various kidney cell lines (Vero, BHK, LLC Mk2) and mosquito cell lines (C6/36 Aedes albopictus API-61) are used. Infected cell lines with cytopathic effect (CPE) or plaque formation can be confirmed by antigen detection or RT-PCR.
Virus isolation, however, is time consuming, labour intensive and also less sensitive than RNA detection by molecular techniques. In addition, it may require additional biosafety level. However, this is useful for characterization of the virus strain.
Serology: Detection of YF virus specific IgM and IgG antibodies can help in confirming the infection.
Confirmation of suspected YF case is made by: (i) Detection of YFV specific IgM antibody in a suspected case without vaccination in previous 30 days, (ii) demonstration of four fold rise in YFV specific IgG antibody in acute and convalescent sera, (iii) detection of neutralizing antibodies by plaque reduction neutralization test (PRNT).
ELISA and indirect immunofluorescence methods are commonly used methods for detection of IgM and IgG antibodies in serum. However, unlike other flaviviruses, reagents are not widely available commercially. CDC has developed and validated an MAC ELISA test for YF IgM antibody detection.
YF IgM antibody mostly appears during the period of remission and peaks during the period of intoxication and persists for a long time. In YF naïve vaccinated individuals, IgM persists for longer period (years) as compared to vaccinated individual from YF endemic regions.
Other serological methods like complement fixation test (CFT), hemagglutination inhibition test (HAI) and plaque reduction neutralization test (PRNT) can be used to detect fourfold rise in antibody titer. However, due to the technical difficulties, these tests are no more in use for patient diagnosis.
Cross-reactivity with other flaviviruses may lead to false positive reaction. Hence, the sample positive for YF serology should be negative for other flaviviruses in order to be considered as specific.
Yellow fever virus RNA detection: Various molecular techniques have been developed for detection of YFV RNA in clinical samples. This includes RT-PCR, real-time PCR, multiplex PCR and isothermal amplification methods. Several commercial PCR assays are available: Single detection or multiplex PCR.
NS1 antigen detection: ELISA for detection of NS1 antigen has been developed and has been shown to be 80% sensitive and 100% specific.
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