Laboratory diagnosis of viral infections
In the diagnostic laboratory virus infections are confirmed by several methods that include:
- Growth of the virus in a cell culture from a specimen taken from the patient.
- Detection of virus-specific antibodies in the blood.
- Detection of virus antigens
- Detection of virus nucleic acids
- Gene sequencing to characterise viral strains
- Observation of virus particles by electron microscopy.
- Hemagglutination assay
Diagnostic virology has changed rapidly due to the advent of molecular techniques and increased clinical sensitivity of serological assays.
A wide variety of samples can be used for virological testing. The type of sample sent to the laboratory often depends on the type of viral infection being diagnosed and the test required. Blood is used for serological testing for antibody and antigen detection in a variety of different infections as well as monitoring viraemias. Sputum, gargles and bronchial washings are used to detect respiratory viruses such as Influenza. Faeces are usually sent to determine the presence of viral gastroenteritis agents. Other samples include swabs (usually in viral transport medium), cerebrospinal fluid, biopsies and post mortem tissues, dried blood spots and others.
Proper sampling technique is essential to avoid potential pre-analytical errors.
When growing virus in a cell culture, the cells affected with virus will evolve morphologic changes, often specific for the type of virus involved.
When the adaptive immune system of a vertebrate encounters a virus, it produces specific antibodies which bind to the virus and render it non-infectious. This is called humoral immunity. Two types of antibodies are important. The first called IgM is highly effective at neutralizing viruses but is only produced by the cells of the immune system for a few weeks. The second, called, IgG is produced indefinitely. The presence of IgM in the blood of the host is used to test for acute infection, whereas IgG indicates an infection sometime in the past. Both types of antibodies are measured when tests for immunity are carried out. Antibody testing has become widely available through automated panels.
Detection of virus antigens can be done by ELISA in tissues and fluids.
Other techniques are:
Many viruses attach to molecules present on the surface of red blood cells. A consequence of this is that - at certain concentrations - a viral suspension may bind together (agglutinate) the red blood cells thus preventing them from settling out of suspension. Usefully, agglutination is rarely linked to infectivity, attenuated viruses can therefore be used in assays.
Nucleic acid detection
Detection of virus encoded DNA and RNA is done with polymerase chain reaction. Nucleic acid hybridization with virus-specific probes detects specific viruses. Molecular techniques are usually used to confirm positive serological results due to their higher sensitivity and specificity. Variations of PCR such as nested reverse transcriptase PCR and real time PCR can also be used to determine viral loads in patient serum. This is often used to monitor treatment success in HIV cases.
Some viruses require specialised gene sequencing techniques to be carried out in order to help predict the type of infection that will display in the patient. Certain genotypes of the same viruses can lead to different transmission routes, virulence and treatment. In some cases, specific mutations in the patient are tested for in order to determine antiviral therapy and susceptibility to infection.