Laboratory Diagnosis

Virus Isolation

Dengue and yellow fever are best diagnosed by virus isolation from blood taken as early as possible during the course of the disease. This is particularly relevant to a suspected indicator case in a region that experiences occasional epidemics of dengue, since it is important to characterize the serotype causing the outbreak. However, other flaviviruses are not readily isolated from human patients, as viremia is frequently brief or undetectable. Postmortem, virus can sometimes be recovered from appropriate organs, such as the brain in cases of encephalitis and the liver in yellow fever. Most arboviruses have been isolated from collections of mosquitoes or other vectors, traditionally by intracerebral injection of newborn mice but more recently by inoculation of cultured mosquito or vertebrate cells. The cytopathic effect varies with the virus and cell type: some combinations produce syncytia, but many give undramatic cyto-pathology or none at all. Viral antigen can be detected by immunofluorescence.

Characterization of isolates relies on serology, combined with a knowledge of what viruses are likely to occur in particular geographic regions and ecosystems. Whereas hemagglutination inhibition is useful for assigning an arbovirus to a particular genus or group, individual flaviviruses display extensive cross-reactivity. Neutralization is the appropriate test for characterization at the species level. Even those viruses that induce no cytopathic effect in cultured cells usually produce plaques; hence, plaque reduction is the most convenient form of neutralization test.

Because of the pronounced cross-reactions between species, monoclonal antibodies have proved to be of great value in research and may become standard diagnostic reagents, particularly for viruses or geographic regions where serological cross-reactions using polyclonal antisera are a problem. For example, one monoclonal antibody might recognize an antigenic determinant common to all flaviviruses, while another recognizes a determinant common to all members of the dengue complex (dengue types 1, 2, 3, and 4), yet another recognizes only a type-specific determinant on dengue type 3, and another distinguishes subtypes of dengue 3. Oligonucleotide maps of the viral RNA discern even finer differences between isolates from different parts of the world and can be very informative in tracing their origins. For instance, such RNA fingerprints distinguish topotypes of dengue type 2 virus from various parts of the world.

Serology

Diagnosis is usually made by demonstrating a rising titer of antibody in the patient's serum. IgM serology, using EIA with anti-human IgM as capture antibody, is ideally suited to rapid diagnosis of individual patients during an epidemic, but it is not sufficiently definitive to rely on for identifying an "indicator" case prior to such an epidemic. However, the reagents used in such tests must be absolutely reliable, and IgM antibodies can persist for many months and/or be elicited by infection with heterologous viruses. Indicator cases should always be diagnosed by rising antibody titer or, where possible, by virus isolation.

Was this article helpful?

0 0

Post a comment