Laboratory diagnosis of viral diseases: Five common approaches

The diagnosis of a viral infection can be established in several ways, often used together, namely:

1. Detection of virus particles in specimen taken from the appropriate site.
2. Detection of viral antigens (Ag) in blood or body fluids
3. Serological procedure to detect specific anti-viral antibodies (rise in antibody titre or presence of IgM antibody)/detection of presence of a cell-mediated immune response.
4. Detection of viral nucleic acids in the blood or body cells of a patient
5. Culture of infectious virus from appropriate clinical specimen.
6. Cytological or histological examination of cells from the site of the infection in those viral infections in which a characteristic viral cytopathic effect (CPE) is produced.

Microscopic identification of Viruses

Viruses can be detected and identified by direct microscopic examination of clinical specimens such as biopsy materials or skin lesions. Three different microscopy techniques are currently in use:

• Light microscopy: It reveals characteristic inclusion bodies or multinucleated giant cells. e.g. Tzanck smear which shows herpesvirus induced multinucleated giant cells (MGCs) in vesicular skin lesions.
• UV Microscopy: It is used for fluorescent antibody staining of the virus in infected cells.
• Electron Microscopy: It detects virus particles, which are further characterized by their size and morphology.

Serological procedure for the laboratory diagnosis of Viruses

• A rise in antibody titre to the virus can be used to diagnose viral infection.  A serum sample is obtained in the acute phase (as soon as viral etiology is suspected), and a second sample is obtained in the convalescent phase (10-14 days later). If the antibody titre in the convalescent phase serum sample is at least 4 fold higher than the titre in the acute phase serum sample, the patient is considered to be infected.
• In some viral disease for which cutoff titre is known, patient showing rise in antibody titre than cut off value can be considered as infected and
• in other viral disease the presence of IgM antibody is the diagnostic. Primary antibody response is governed by IgM (predominately produced).

Detection of Viral Antigens

Various tests such as ELISA, EFA etc can be done to detect the presence of viral antigens in the patient blood or biopsy materials. For the diagnosis of Hepatitis virus infection, HBsAg (Hepatitis viral surface antigen) or HBeAg (Hepatitis virus e antigens) can be detected. Similarly, detection of p24 viral antigen is the diagnostic method in case of HIV Infection.

Detection of Viral Nucleic acids

Detection of viral nucleic acids is one of the sensitive and rapid methods for laboratory diagnosis. It requires the use of PCR (polymerase chain reaction) to amplify the viral genome present in the sample and detection of the specific gene sequence of that particular virus by the use of a specific primer (while performing PCR) and probe (while detecting the specific sequence).  RNA viral assay is currently in use to monitor the course of HIV infection and to evaluate the patient progress.

Identification of the virus in the cell culture

Viruses are obligate intracellular parasites so we cannot grow them in ordinary media (like we grow bacteria, fungi) and require living cells for the growth and propagation.  The growth of virus in the cell culture may produce characteristic cytopathic Effect ( CPE) which helps us for presumptive diagnosis.  If that particular virus does not produce the cytopathic effect, its presence can be detected by several other techniques such as Immunofluorescence assay (eg. DFA, IFA), Radioimmunoassay (RIA), Hemadsorption, decrease in acid production of infected cells, ELISA, Complement fixation, Hemagglutination inhibition method, Neutralization etc.